REVIEWS

New aspects of vitamin D metabolism

and action — addressing the skin as
source and target
Daniel Bikle   1,2* and Sylvia Christakos3
Abstract | Vitamin D has a key role in stimulating calcium absorption from the gut and promoting
skeletal health, as well as many other important physiological functions. Vitamin D is produced in
the skin. It is subsequently metabolized to its hormonally active form, 1,25-dihydroxyvitamin D
(1,25(OH)2D), by the 1-hydroxylase and catabolized by the 24-hydroxylase. In this Review , we pay
special attention to the effect of mutations in these enzymes and their clinical manifestations. We
then discuss the role of vitamin D binding protein in transporting vitamin D and its metabolites from
their source to their targets, the free hormone hypothesis for cell entry and HSP70 for intracellular
transport. This is followed by discussion of the vitamin D receptor (VDR) that mediates the cellular
actions of 1,25(OH)2D. Cell-specific recruitment of co-regulatory complexes by liganded VDR
leads to changes in gene expression that result in distinct physiological actions by 1,25(OH)2D,
which are disrupted by mutations in the VDR . We then discuss the epidermis and hair follicle, to
provide a non-skeletal example of a tissue that expresses VDR that not only makes vitamin D but
also can metabolize it to its hormonally active form. This enables vitamin D to regulate epidermal
differentiation and hair follicle cycling and, in so doing, to promote barrier function, wound healing
and hair growth, while limiting cancer development.

1
Departments of Medicine
and Dermatology, University
of California San Francisco,
San Francisco, CA, USA.
2
VA Medical Center,
San Francisco, CA, USA.
3
Departments of
Microbiology, Biochemistry
and Molecular Genetics,
New Jersey Medical School,
Rutgers, the State University
of New Jersey, Newark,
NJ, USA.
*e-mail: Daniel.bikle@
ucsf.edu
https://doi.org/10.1038/
s41574-019-0312-5
Vitamin D, a fat-soluble secosteroid, comes in two forms:
vitamin D2 and vitamin D3. These forms differ only in the
side chain, where vitamin D2 has a double bond between
C22 and C23, and a methyl group at C24, unlike vitamin
D3. Vitamin D3 is produced in the skin from 7-dehydro-
cholesterol (7-DHC)1, whereas vitamin D2 is produced in
plants and fungi from ergosterol2. The biological activi-
ties are comparable, although there are subtle differences
in pharmacokinetics3; unless stated otherwise in the text,
vitamin D with no subscript refers to both forms.
Vitamin D3 is produced in the skin via a two-step pro-
cess under the influence of ultraviolet B (UVB) radia-
tion (290–315 nm) from the vitamin D substrate 7-DHC.
The synthesis begins with the conversion of 7-DHC to
pre-vitamin D, which then undergoes thermal conversion
to vitamin D4. Clothing and deep pigmentation of skin
limit epidermal vitamin D production, but, according to
a 2019 review, sunscreens do not limit production very
much5. The conversion of 7-DHC to pre-vitamin D and
subsequent conversion of pre-vitamin D to vitamin D
are non-enzymatic steps. The production 7-DHC, how-
ever, is enzymatic. 7-DHC is produced in the skin via the
Kandutsch–Russell pathway. This pathway results in
the production of either cholesterol by the enzyme 7-DHC
reductase (DHCR7) or vitamin D by UVB radiation6.
The Kandutsch–Russell pathway is an alternative path-
way to the Bloch pathway for cholesterol synthesis that
directs zymosterol to 7-DHC, which can be converted
by UVB radiation to vitamin D, rather than to des-
mosterol, which cannot. Conversion to desmosterol is
achieved by reversing the order in which 24-dehydro-
cholesterol reductase (DHCR24) and DHCR7 act at the
different stages of cholesterol synthesis. The Bloch path-
way is primarily utilized by steroidogenic tissues, liver
and kidney, whereas the Kandutsch–Russell pathway is
primarily utilized by skin, brain and muscle6. Therefore,
DHCR7 is really the critical first step in the production of
vitamin D, and, as discussed below, its regulation impacts
the amount of 7-DHC available for photoproduction of
vitamin D in the skin.
Vitamin D has limited biological activity, although it
does inhibit Smoothened, which is a key component of
the Hedgehog pathway and might be part of the means
by which vitamin D signalling protects the skin from the
carcinogenic effect of UVB radiation7. For most vitamin
D-mediated actions, vitamin D must first be metabolized
to 25-hydroxyvitamin D (25OHD), a process that occurs
primarily, but not exclusively, in the liver8. 25OHD is the
major circulating metabolite of vitamin D9. Numerous
enzymes have 25-hydroxylase activity, but the major

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Key points
• Mutations in the enzymes involved in vitamin D metabolism cause human diseases
such as rickets.
• Vitamin D binding protein is the major transport protein for vitamin D metabolites,
but with the exception of a few tissues such as the kidney, it is the free forms of these
metabolites that enter cells.
• The vitamin D receptor (VDR) is the major mediator of vitamin D biological action.
• VDR binding sites can be located in a range of locations including introns and at distal
intergenic regions of regulated genes.
• Co-regulators of VDR provide cell-specific genomic regulation.
• The skin is an excellent non-skeletal model for the study of vitamin D as it is the source
of vitamin D, contains the vitamin D-metabolizing enzymes and expresses the VDR,
making it a target tissue as well.
enzyme responsible for 25OHD production under phys-
iological circumstances is CYP2R1. Knockout of Cyp2r1
in mice results in a reduction in blood levels of 25OHD by
>50%, but not to zero10. Moreover, mutations in CYP2R1
in humans leads to rickets11. Although 25OHD is the
major circulating form of vitamin D, its active hormo-
nal form, which is synthesized by the enzyme CYP27B1,
is 1,25(OH)2D.
Renal cells are the major source of circulating
1,25(OH)2D, but many other cells, including keratino-
cytes and immune cells, express CYP27B1. The regula-
tion of the production of 1,25(OH)2D in non-renal cells
differs from that in renal cells, leading to the concept that
non-renal cells are capable of producing 1,25(OH)2D
for their own needs in an autocrine and/or paracrine
fashion12. If the activity of CYP27B1 is unchecked, the
result can be the over-production of 1,25(OH)2D, which
can result in hypercalcaemia and/or hypercalciuria12.
In the kidney, CYP27B1 activity is stimulated by para-
thyroid hormone (PTH) and inhibited by the action of
fibroblast growth factor 23 (FGF23) and 1,25(OH)2D.
In other tissues, the regulation of CYP27B1 occurs
primarily by cytokines such as tumour necrosis factor
and interferon-γ12. 1,25(OH)2D indirectly regulates its
own levels in these cells via CYP24A1, a 24-hydroxy-
lase. CYP24A1, which is found in most tissues, is key
to the catabolism of both 25OHD and 1,25(OH)2D. As
the production of CYP24A1 is induced by 1,25(OH)2D,
CYP24A1 serves as the brake on levels of 1,25(OH)2D.
Not surprisingly, mutations in CYP27B1 and CYP24A1
cause profound disturbances. Inactivating mutations in
CYP2R1 and CYP27B1 lead to rickets13,14, and inactivat-
ing mutations in CYP24A1 are one cause of idiopathic
infantile hypercalcaemia (IIH) in children and recurrent
kidney stone formation in adults15.
Vitamin D and its metabolites are generally trans-
ported from the site of production to their target tis-
sues. Vitamin D binding protein (DBP), which binds
to ~85% of the circulating 25OHD and 1,25(OH)2D,
is primarily responsible for such transport16,17. DBP is
a highly polymorphic protein, and polymorphisms
can affect its functions, including its ability to bind to
25OHD18. The finding that polymorphisms in DBP can
alter its function has sparked new interest in defining
the poly­morphisms, the effect they have on circulating
levels of vitamin D and the response of individuals to
vitamin D supplementation with respect to increases in
total and free 25OHD19. However, with the exception of
cells (such as renal cells) that express the megalin–cubilin
complex, most cells are thought to take up 25OHD and
1,25(OH)2D in the non-protein-bound form, a concept
termed the free hormone hypothesis20. This hypothesis
has generated a discussion as to whether free 25OHD
might provide a better measure of vitamin D status than
total 25OHD. In this regard, studies in mice lacking
DBP and a case report of a patient lacking DBP indicate
that loss of DBP does not result in vitamin D deficiency,
even in the presence of undetectable 25OHD levels
unless intake of vitamin D is restricted21,22.
Within the cells that have been studied, HSP70 has
been identified as a potential transporter of 25OHD
through the plasma membrane to the mitochon-
dria, where it is converted to 1,25(OH)2D23. From the
mitochondria, HSP70 is then thought to transport
1,25(OH)2D into the nucleus where it binds to the vita-
min D receptor (VDR)23. HSP70 has a higher affinity
for 25OHD than DBP, which facilities 25OHD trans-
port across the cell membrane, but lower affinity for
1,25(OH)2D than VDR, which facilities 1,25(OH)2D
offloading onto VDR in the nucleus23.
VDR also mediates the genomic, and some of the
non-genomic actions, of 1,25(OH)2D24,25. In addition,
VDR has actions that are independent of 1,25(OH)2D,
including the regulation of hair follicle cycling 26.
Furthermore, each cell has its own repertoire of responses
to 1,25(OH)2D, indicating that within different cells are
factors, such as co-regulators and other transcription fac-
tors, that influence VDR function in a cell-specific man-
ner. One excellent example of the cell-specific actions of
1,25(OH)2D and/or VDR is the regulation of Cyp27b1 in
renal cells in contrast to the lack of regualtion of Cyp27b1
activity by 1,25(OH)2D/VDR in macrophages27. In par-
ticular, the regulatory region of Cyp27b1 in mouse renal
cells that responds to 1,25(OH)2D is not responsive to
1,25(OH)2D in macrophages and most other non-renal
cells27. Over the past 5 years, genetic and epigenetic
mechanisms have been discovered that underlie the
tissue-specific actions of VDR and its ligand. The differ-
ent regulation of CYP27B1 and CYP24A1 by hormones
and cytokines between renal cells and non-renal cells
is a good example of this that is relevant to vitamin D
metabolism27,28, as is the different regulation of receptor
activator of nuclear factor-κΒ ligand (RANKL) by hor-
mones and cytokines between osteoblasts and T cells29.
Furthermore, structural analyses have examined how
mutations in the VDR can affect its function, including
its interactions with the various co-regulators, with some
mutations in the zinc finger and hinge region interfering
with retinoid X receptor (RXR) binding and others in
the AP2 domain interfering with co-activator binding30.
The skin is the only organ that has the capacity to
make vitamin D, metabolize it to 1,25(OH)2D, and res­
pond to it in a cell-specific fashion31. Psoriasis, a prolif-
erative inflammatory disease of the skin, is a pro­minent
example of a non-skeletal disease in which 1,25(OH)2D
and its analogues have proved to be effective ther-
apy32. This finding illustrates both the antiproliferative
and anti-inflammatory actions of vitamin D. The cells
responsible for these responses to 1,25(OH)2D in the
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skin are keratinocytes, the dominant cell type of both
the epidermis and hair follicles1. As such, the skin pro-
vides an excellent model tissue in which to study both
vitamin D metabolism and its mechanisms of action,
as the regulation of metabolism in keratinocytes differs
from that in the kidney31. For these reasons, the free
hormone hypothesis can be directly tested in the skin.
Furthermore, in the skin the cell-specific actions of VDR
and/or 1,25(OH)2D are clearly illustrated by the differ-
ent molecular events that are regulated by VDR and/or
1,25(OH)2D during the temporal and sequential events
of epidermal differentiation and hair follicle cycling1.
In this Review, we discuss what we have learned from
the new discoveries in vitamin D metabolism. We focus
on mutations in the enzymes that metabolize vitamin D,
DBP and its role in regulating tissue access to the
vitamin D metabolites it transports, and the mechanisms
that underlie the tissue-specific actions of VDR. In the
final section we use the skin to illustrate a number of
these points, with a focus on the diversity of vitamin D
actions within the vitamin D endocrine system, and
how this diversity is utilized in the regulation of three
important functions of the skin; namely, protection
from the environment, promotion of wound healing and
suppression of cancer development.
Vitamin D production and metabolism
In this section, we discuss the key enzymes involved in
vitamin D production and metabolism including the
impact of mutations in these enzymes. Figure 1 shows the
overall pathway that is discussed, and Box 1 summarizes
the mutations and their clinical features.
DHCR7. As noted in the introduction, DHCR7 regu-
lates the levels of 7-DHC in the skin. Its activity con-
verts 7-DHC to cholesterol. When DHCR7 is inhibited
(or inactivated by mutations), more 7-DHC is available
for vitamin D production. In addition to the skin,
this enzyme is found in the brain, muscle and heart33.
Cholesterol and vitamin D (but not 1,25(OH)2D) incre­
ase proteasomal degradation of DHCR7, leading to
increased vitamin D production34. AMP-activated pro-
tein kinase, a key sensor and regulator of cellular energy
homeostasis, and protein kinase A, are potent inhibitors
of DHCR7 (refs6,35).
Inactivating mutations of DHCR7 result in the
Smith–Lemli–Opitz syndrome, a developmental dis­
order in which patients can present with mental
retardation, growth deficiency and congenital malfor-
mations36,37. These patients are affected primarily by the
consequences of too little cholesterol, steroids or bile
acids. However, they seem to be more sensitive to UVB
radiation, and can present with increased serum 25OHD
concentrations 38, although not with toxic levels 38.
Polymorphisms in DHCR7 have been associated with
either reduced39,40 or increased41 25OHD levels. Whether
these polymorphisms, all of which are in introns, affect
enzymatic function is not clear.

↑ P and Ca OH
OH
↑ 1,25(OH)2D
↑ FGF23
↓ PTH

CH2
OH CYP24A1
HO
24,25(OH)2D3
Liver Kidney

CYP2R1
CH2 CH2
CYP27B1
OH
HO HO
D3 25OHD3
↓ P and Ca
↑ PTH
↓ FGF23 CH2

HO OH
1,25(OH)2D3

Fig. 1 | Vitamin d production and metabolism. 7-Dehydrocholesterol reductase (DHCR7) is involved in the last step in
cholesterol synthesis, which in the case of the skin competes with vitamin D production as each requires the substrate
7-dehydrocholesterol (7-DHC). Thus DHCR7 activity is relevant to vitamin D production as it controls the level of the
substrate for vitamin D production, namely 7-DHC. When exposed to UVB radiation, the path to vitamin D in the skin is
initiated with breaking of the B ring of 7-DHC to form previtamin D3, which undergoes isomerization to vitamin D3. Vitamin D3
then undergoes two hydroxylations first in the 25 position principally in the liver by CYP2R1 and then in the kidney either
to 24(OH)2D3 by CYP24A1 or to the active metabolite 1,25(OH)2D3. In the kidney, these latter hydroxylations are tightly
controlled by parathyroid hormone (PTH) that stimulates CYP27B1 and inhibits CYP24A1 and fibroblast growth factor 23
(FGF23) that does the reverse. 1,25(OH)2D3 induces CYP24A1 while suppressing CYP27B1. Calcium and phosphate directly
or via their regulation of PTH and FGF23 production likewise participate in this regulation. However, in extrarenal tissues,
regulation of CYP27B1 is different, and depends primarily on various cytokines functioning as autocrine or paracrine factors.

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Cerebrotendinous
CYP2R1 and other 25-hydroxylases. As mentioned in the
xanthomatosis Introduction, the liver is the major, if not the sole, source of
The disease that results from 25OHD production from vitamin D8. 25-Hydroxylation
inactivating mutations in activity in the liver has been detected in both the mito-
CYP27A1. Cerebrotendinous
chondrial and microsomal fractions 8. Numerous
xanthomatosis manifests as a
reduction in bile and enzymes, including CYP3A4 and CYP27A1, have sub-
cholesterol metabolism. sequently been found to have 25-hydroxylase activity42.
Initial studies suggested that CYP27A1, a mitochondrial
enzyme with substantial homology to both CYP27B1 and
CYP24A1 (the 1α and 24-hydroxylases, respectively),
was the major 25-hydroxylase; however, several prob-
lems subsequently emerged with this identification. For
example, CYP27A1 was originally identified as a sterol
27-hydroxy­lase involved in bile acid synthesis, and inacti­
vating mutations result in cerebrotendinous xanthomatosis
with abnormal bile and cholesterol metabolism, but not
rickets43. Moreover, CYP27A1 does not 25-hydroxylate
vitamin D2 (refs2,3). Furthermore, in mouse studies,
deletion of Cyp27a1 resulted in increased blood levels of
25OHD rather than a decrease, indicating that expres-
sion of CYP27A1 is not required for the production of
25OHD, and might even be inhibitory10.
Current data support CYP2R1 as the major
25-hydroxylase in the liver. This enzyme 25-hydroxylates
both vitamin D 2 and vitamin D 3 equally, unlike
CYP27A1, which does not hydroxylate vitamin D2 as
noted above. In humans, CYP2R1 is expressed primar-
ily in the microsomal fraction of the liver and testes42,44.
As noted in the Introduction, deletion of Cyp2r1 in
mice reduces but does not eliminate 25OHD produc-
tion and has little effect on blood levels of calcium and
phosphate10. These findings indicate the presence of
a compensatory mechanism by other enzymes with
25-hydroxylase activity. Even the double deletion of
Cyp2r1 and Cyp27a1 in mice does not reduce the blood
level of 25OHD to zero10. In humans, however, mutations
in CYP2R1 have a greater inhibitory effect on 25OHD
levels and result in rickets, but the severity of the rickets
is not as great as seen with mutations in CYP27B1 (ref.13).
Box 1 | Mutations in the vitamin d metabolism pathway
• 7-dehydrocholesterol reductase (dHcr7). This enzyme converts the substrate also serves to degrade both 25OHD and 1,25(OH)2D
for vitamin D formation (7-dehydrocholesterol) to cholesterol. Polymorphisms are
associated with reduced 25-hydroxyvitamin D (25OHD) levels, and inactivating
mutations have been associated with increased vitamin D production38.
• cyP2r1. This enzyme is considered the major vitamin D 25-hydroxylase. Inactivating
mutations result in a form of rickets currently called vitamin D-dependent rickets,
type 1B (VDDR1B)46. Affected individuals have low 25OHD levels but normal
1,25-dihydroxyvitamin D (1,25(OH)2D) levels. They can be treated with 25OHD
(calcidiol) and may no longer require treatment as adults.
• cyP27B1. This enzyme is the 25OHD-1α hydroxylase responsible for producing the
hormonal form of vitamin D — 1,25(OH)2D. Mutations in this enzyme result in a form
of rickets called pseudovitamin D deficiency rickets or vitamin D-dependent rickets,
type 1A (VDDR1A)52. Affected individuals present with low 1,25(OH)2D but normal
levels of 25OHD. They respond to 1,25(OH)2D (calcitriol) but not to vitamin D.
• cyP24A1. This enzyme is the 24-hydroxylase with both 25OHD and 1,25(OH)2D as
substrates. Its primary role is that of catabolizing its substrates to inactive forms.
Mutations result in a form of idiopathic infantile hypercalcaemia (IIH), although with
the discovery of these mutations, this form of IIH is no longer idiopathic and, as the
disease can manifest in adults, is no longer just infantile15,63. In children, hypercalcaemia
is the major concern. Presentation in adults is usually with a history of kidney stones
associated with hypercalcaemia and elevated 1,25(OH)2D levels.
To date, five functional mutations in CYP2R1 have
been described. The first was reported in a young man
from Nigeria who had severe bone disease and bio-
chemical evidence of rickets, including a low 25OHD
level but a normal 1,25(OH)2D level. This individual
had a Leu99Pro mutation in CYP2R1, the first mutation
identified11. In vitro testing confirmed that this muta-
tion decreased the activity of the mutated CYP2R1 to
produce 25OHD11. Subsequently, an additional muta-
tion, Lys242Asn, was found in an individual carrying
compound heterozygote mutations in CYP2R1, when
additional Nigerian families with familial rickets were
screened13. Two more mutations were found in siblings
of an Arabian family carrying compound heterozygote
mutations involving a splice site (367+1, Gly to Ala) and
an insert (768, iThr)45. Finally a mutation involving a
deletion — Gly42_Leu46 with an inserted Arg — was
found in a French family46. These mutations, when
tested in vitro, showed little or no 25-hydroxylase activ-
ity13,45,46. In addition, the identified individuals main-
tained normal or even high 1,25(OH)2D levels, and some
responded to treatment with vitamin D and 1αOHD,
with a further increase in 1,25(OH)2D following treat-
ment13,46. Moreover, as the individuals reach adulthood,
they generally maintained normal calcium homeosta-
sis without vitamin D supplementation46. Thus, other
enzymes with 25-hydroxylase activity might be upregu­
lated to compensate for the reduced CYP2R1 activity.
Nevertheless, the affected individuals developed classic
rickets, albeit sometimes not diagnosed until early teens,
along with high PTH and alkaline phosphatase levels as
well as very low 25OHD levels13,45,46. Affected individu-
als responded very well to physiological doses of calci­
fediol (25OHD)13. This form of rickets, comparable to
vitamin D deficiency rickets with disrupted endochon-
dral bone formation and impaired mineralization of oste-
oid affecting growth and skeletal integrity, is currently
called vitamin D-dependent rickets type 1B (VDDR1B).
One of the aforementioned compensating enzymes
could be CYP3A4, the major drug-metabolizing enzyme
that is preferentially located in the liver and intestine47.
This enzyme has 25-hydroxylase activity47, however, it
with hydroxylations in the 23 and 24 positions as well,
in a similar manner to CYP24A1 (ref.48). An activating
mutation in CYP3A was described in 2018 that increases
the catabolism of 1,25(OH)2D and presumably 25OHD
leading to rickets with reductions in both 25OHD and
1,25(OH)2D, decreased serum calcium and phosphate
with elevated PTH and alkaline phosphatase. The
authors of this study called this type of rickets vitamin
D-dependent rickets type 3 (ref.49).
CYP27B1. To date, CYP27B1 is the only 25OHD-1α
hydroxylase to have been described. Within cells, this
enzyme is found in mitochondria, and while the kidney
might be the main contributor to circulating 1,25(OH)2D,
a number of other tissues express this enzyme includ-
ing the epidermis and other epithelial tissues, bone, pla-
centa and cells of the immune system12. The regulation
of CYP27B1 expression in these sites outside the kid-
ney differs from that in the kidney, where regulation by
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PTH and FGF23 dominate12. In non-renal tissues,
CYP27B1 is regulated by a number of cytokines including
interferon-γ and tumour necrosis factor, but not by cal-
cium, PTH or 1,25(OH)2D50. This accounts for the hyper-
calcaemia with inappropriate elevations of 1,25(OH)2D
that complicate a number of granulomatous diseases
and lymphomas, demonstrating stimulation by these
cytokines overexpressed in these diseases with lack of
feedback by the elevation of calcium and 1,25(OH)2D and
suppressed PTH12,51. In this section, we focus specifically
on the mutations in CYP27B1.
Mutations in CYP27B1 cause a disease previously
referred to as pseudovitamin D deficiency rickets,
which is now known as vitamin D-dependent rickets
type 1A (VDDR1A). A few years after the discovery
of 1,25(OH)2D as the active metabolite of vitamin D
and the development of assays to measure it, Fraser and
colleagues52 and Scriver and colleagues53 were the first
to describe patients with VDDR1A. The investiga-
tors observed that 1,25(OH)2D levels were very low in
patients who presented with rickets despite normal lev-
els of 25OHD, distinguishing them from other patients
with vitamin D-deficient rickets. They called the dis-
ease pseudovitamin D deficiency rickets because they
could not cure the rickets with vitamin D. Several years
after the first descriptions of VDDR1A, Delvin and col-
leagues54 found that VDDR1A can be treated effectively
with 1,25(OH)2D (calcitriol), but not with vitamin D.
It was not until 1997, however, that the cloning and
sequencing CYP27B1 was accomplished14,55–57. In one
of these studies, Portale and colleagues sequenced the
human gene for the first time and identified the first mut­
ation in a patient with VDDR1A14. Subsequently, muta-
tions have been found throughout the gene58. Both the
renal and extrarenal CYP27B1 have the same sequence,
but their differences in regulation occur as a result
of tissue-specific multicomponent control modules,
as noted in a 2017 article describing loss of function
enhancer deletion studies that demonstrated different
regions of the CYP27B1 promoter region responding to
different regulators in different cells27.
The clinical symptoms of rickets generally occur in
the first year of life — embryonic development is nor-
mal52. In addition to the skeletal defects of classic rickets
(flared metaphyses, bowing of the legs, rachitic rosary
and generalized osteopenia), patients develop tooth
dysplasia, muscle weakness and growth retardation,
and can have convulsions and/or tetany due to the very
low calcium levels59. Pseudofractures can develop in the
long bones59. Laboratory assessment in addition to low
1,25(OH)2D include hypocalcaemia, hypophosphatae-
mia and secondary hyperparathyroidism. In patients
with rickets, 25OHD levels are generally normal or even
elevated when dietary insufficiency is excluded59. Thorax
deformities can lead to poor air exchange with predis-
position to pulmonary infections59. As mentioned previ-
ously, physiological doses of calcitriol (1,25(OH)2D) are
curative59. Distinguishing rickets from vitamin D defi-
ciency, especially in countries where low calcium and
vitamin D intake are common, can be difficult59. As such,
a careful family and dietary history is necessary; how-
ever, it is important to note that patients with vitamin D
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deficiency present with low 25OHD levels and often
normal 1,25(OH)2D levels, except those with extreme
vitamin D deficiency59. Patients with hypophosphatae-
mic rickets (such as X-linked or autosomal dominant
hypophosphataemic rickets) will also present with low
1,25(OH)2D and low phosphate levels, but these forms
of rickets are associated with elevated FGF23 levels,
which is not found in those with CYP27B1 mutations60.
As noted above, calcitriol is the treatment of choice, but
needs to be accompanied by an adequate intake of cal-
cium59. Monitoring urine calcium excretion is necessary
to avoid overdosing that can result in nephrocalcinosis
and stone formation59.
CYP24A1 and other 23- and 24-hydroxylases. In most tis-
sues, CYP24A1 is the dominant 24-hydroxylase involved
in vitamin D metabolism. The liver and intestine seem
to be exceptions in that CYP3A4 is highly expressed
in these tissues, although CYP3A4 lacks the specificity
for vitamin D metabolites shown by CYP24A1 (ref.48).
Nevertheless, its induction by drugs such as rifampin can
result in reduced levels of 25OHD and 1,25(OH)2D, which
leads to osteomalacia61. Both CYP24A1 and CYP3A4 have
24-hydroxylase and 23-hydroxylase activity, although,
at least for CYP24A1, the ratio is species-specific in that in
humans 24-hydroxylase activity predominates, whereas
in the opossum 23-hydroxylase predominates62,63. Both
enzymes are induced by 1,25(OH)2D, and in the intestine
the induction of CYP3A4 seems to be at least as great as
that of CYP24A1 (ref.64). That said, CYP24A1 is the domi-
nant vitamin D catabolic enzyme in most tissues and thus
is the focus of the remainder of this section.
The 24-hydroxylase pathway terminates with the bio-
logically inactive calcitroic acid, whereas the 23-hydroxy­
lase pathway produces the biologically active 1,25,26,23
lactone. CYP24A1 catalyses all steps in these pathways65.
Both 1,25(OH)2D and 25OHD are metabolized by
CYP24A1, although 1,25(OH)2D is the preferred sub-
strate63. CYP24A1 is generally considered a catabolic
enzyme in vitamin D metabolism, but this is only par-
tially true as some of its products have biological activ-
ity. For example 1,24,25(OH)3D has substantial affinity
for the VDR with biological activity being ~10% that
of 1,25(OH)2D66. A specific receptor for 24,25(OH)2D
has been identified in bone and other tissues, such as
the skin. This receptor has been shown to be involved
in fracture repair67 and might have a role in mediating
the effects of 24,25(OH)2D on endochondral bone for-
mation68. The deletion of CYP24A1 in mice, however,
resulted in defective mineralization of intramembranous
(not endochondral) bone69, although this seemed to be
due to the elevated levels of 1,25(OH)2D in the knockout
mouse and not to the loss of 24,25(OH)2D per se.
Inactivating mutations in CYP24A1 have been estab-
lished as a major cause of idiopathic infantile hypercalce-
mia (IIH), a syndrome that is associated with severe
hypercalcaemia, hypercalciuria and nephrocalcinosis,
decreased PTH, low 24,25(OH)2D and inappropriately
normal-to-high 1,25(OH)2D15. In a 2017 survey, 21 mis-
sense mutations were reported in the CYP24A1 gene70.
With regards to the clinical presentation of patients,
not all patients with mutations in CYP24A1 present
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a Free hormone hypothesis b Megalin–cubilin-mediated uptake
Free 25OHD
DBP DBP
25OHD
Megalin Cubilin
Mitochondrion
HSP70 DBP
HSP70 VDR
1,25(OH)2D Nucleus
Intracellular HSP70 shuttle
Fig. 2 | Transport of 25oHd between and within cells. Vitamin D binding protein (DBP)
is the major transport protein for the vitamin D metabolites within the bloodstream.
For most cells, however, it is the unbound or free form of 25-hydroxyvitamin D (25OHD) that
enters cells, whereas in a few tissues such as the kidney and probably also the parathyroid
gland and placenta, DBP with its bound 25OHD enters the cell via the megalin–cubilin
complex. Once in the cell, HSP70 functions as an intracellular shuttle first offloading
25OHD from DBP, moving it to mitochondria for metabolism to 1,25(OH)2D, and subsequently
facilitating the movement of 1,25(OH)2D into the nucleus for binding to the vitamin D
receptor (VDR). The generality of this intracellular shuttle mechanism has not been
established for all target cells.
in childhood. For example, several patients presented
during pregnancy, presumably because of the increased
production of 1,25(OH)2D by the placenta71,72. These
patients had a history of renal stones. Other adults
with this syndrome have been described who were not
pregnant73,74. Such adults generally present with early
onset nephrolithiasis and/or nephrocalcinosis, and the
hypercalcaemia is generally less severe than in those
presenting as infants. Although CYP24A1 mutations
are generally recessive, adults and several infants hetero­
zygous for CYP24A1 mutations who presented with IIH
or nephrocalcinosis have been described74,75. These indi-
viduals typically came from families with a history of
renal stones, indicating that screening such families for
CYP24A1 mutations may be warranted.
IIH was first described in the UK in the 1950s
when infants were routinely treated with high doses of
vitamin D to prevent rickets76. The exact cause of IIH,
however, was not known at that time. We now know that
IIH can be caused by at least two other conditions from
which patients with CYP24A1 mutations should be dis-
tinguished. The first is the Williams–Beuren syndrome.
Children with Williams–Beuren syndrome also present
with hypercalcaemia during infancy, but have a number of
phenotypic features that distinguish them from children
with CYP24A1 mutations who have IIH77. For example,
children with Williams–Beuren syndrome have a deletion
in chromosome 7 that includes loss of a transcription fac-
tor, the Williams syndrome transcription factor (WSTF),
which can cause hypercalcaemia due to unchecked VDR
activity. The second is a hypophosphataemic syndrome
resulting from mutations in SLA34A1, which encodes the
renal sodium phosphate transporter A1. Children with
this hypophosphataemic syndrome can also present with
hypercalcaemia with increased 1,25(OH)2D levels78,79.
However, 24,25(OH)2D levels are normal in patients
with this condition79. CYP24A1 mutations can be identi-
fied by determining the ratio of 25OHD to 24,25(OH)2D
by tandem mass spectrometry80 with confirmation of
the findings by gene sequencing80. Vitamin D-sufficient
subjects have a 25OHD:24,25(OH)2D ratio between 5
and 25, whereas patients with CYP24A1 mutations have
a ratio above 80 (ref.80) (Box 1). Treatment involves ces-
sation of all vitamin D supplements and if necessary a
low-calcium diet.
Cellular transport of D metabolites
In this section, we describe the main transport of vita-
min D and its metabolites in blood via DBP, the role
of DBP in regulating the transport of these metabolites
into cells including the free hormone hypothesis, and
the intracellular transport of the vitamin D metabolites
by HSP70 (Fig. 2).
DBP. DBP (originally known as Gc-globulin) was dis-
covered in 1959 (ref.81), but it was not until 1975 that
its function as a carrier of vitamin D metabolites was
described82. In healthy non-pregnant individuals, the
binding of 25OHD and 1,25(OH)2D to DBP accounts
for ~85% of the total levels of these vitamin D metabo-
lites, whereas albumin with its lower affinity binds ~15%
of the total levels of the vitamin D metabolites. The
non-protein-bound or free 25OHD fraction comprises
about 0.03% of the total 25OHD, with free 1,25(OH)2D
slightly higher at 0.4% of the total 1,25(OH)2D. The
affinity of DBP for the vitamin D2 metabolites is less than
the affinity of DBP for 25OHD and 1,25(OH)2D83, such
that the free fraction would be expected to be higher, but
this has not been directly determined.
For lipophilic hormones such as steroid hormones,
thyroid hormone and vitamin D metabolites, it is the free
fraction that is thought to enter cells — a concept known
as the free hormone hypothesis84. However, at least for
some tissues, a transport system has been identified that
takes up the 25OHD (and presumably other vitamin
D metabolites) attached to DBP. This system involves
megalin and/or cubilin85. This complex is found pri-
marily in the kidney, parathyroid gland, placenta and a
few other tissues19, most of which have not been tested
as to the role of this complex in taking up DBP-bound
vitamin D metabolites. However, mice lacking the
megalin and/or cubilin complex have poor survival
with evidence of osteomalacia, which indicates its role
in vitamin D transport into critical cells involved with
vitamin D signalling85. Knockout of Lrp2 (encoding
megalin) or Dbp genes in mice provides a useful tool for
determining the respective roles of DBP and megalin
and/or cubilin in transporting vitamin D metabolites
into cells. In Dbp-knockout mice the vitamin D metab-
olites are presumably all free and/or bioavailable21,86.
Unlike the Lrp2-knockout mice, mice lacking Dbp do
not show evidence of vitamin D deficiency unless placed
on a vitamin D-deficient diet despite having very low
levels of serum 25OHD and 1,25(OH)2D21. In subse-
quent studies, tissue levels of 1,25(OH)2D were found to
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be normal in the Dbp-knockout mice as were markers of
vitamin D action such as expression of intestinal TRPV6,
calbindin 9k (also known as protein S100G), PMCA1b
and renal TRPV5 (ref.86). These studies support the
concept that DBP serves as a critical reservoir for
the vitamin D metabolites reducing the risk of vitamin D
deficiency when epidermal production or intake is low,
but entry of vitamin D metabolites into cells involves the
free fraction in most cells. The more detrimental effect
of deleting the LRP2 gene, however, might be explained
by the inability of the kidney to reclaim the DBP-bound
metabolites that are then lost in the urine85.
In further support of the mouse studies, a 2019 case
report reported a large deletion in the coding portion
of the DBP gene (and adjacent NPFFR2 gene) in a sin-
gle family22. The proband had normal calcium, phos-
phate and PTH levels with vitamin D supplementation
despite very low levels of 25OHD, 24,25(OH)2D and
1,25(OH)2D that were not responsive to massive doses
of vitamin D (oral or parenteral). The free 25OHD
was nearly normal. The carrier sibling had vitamin D
metabolite levels between those of the proband and the
healthy sibling. Thus, studies in both Dbp-null mice and
humans with mutations in DBP support the free hor-
mone hypothesis while also supporting the role of DBP
as a circulating reservoir for vitamin D metabolites.
Expression of DBP is widespread, but by far the great-
est proportion originates in the liver87. The expression of
DBP is increased by oestrogen88, which is illustrated by
the rise in DBP during pregnancy89,90 and with oral con-
traceptive use91. Androgens, on the other hand, do not
seem to affect DBP expression88. Dexamethasone, and
certain cytokines such as IL-6 also increase DBP pro-
duction, whereas transforming growth factor-β (TGFβ)
is inhibitory92. These drugs and cytokines contribute
to the changes in DBP production following trauma93
and acute liver failure94. Primary hyperparathyroidism,
on the other hand, is associated with a reduction in
DBP levels, that is likely to contribute to the reduced
25OHD levels in these patients as the free 25OHD is not
reduced95. Vitamin D does not regulate DBP production,
nor do any of its metabolites92,96.
Human DBP is 58 kDa, although differences in glyco-
sylation of the protein can alter its size87. DBP is the most
polymorphic gene known. Over 120 variants have been
described based on electrophoretic properties97 with
1,242 polymorphisms currently listed in the NCBI data-
base98. Of these variants, the Gc1F and Gc1S (rs7041)
and Gc2 (rs4588) are the most common. The Gc1F and
Gc1S involve two polymorphisms, one at amino acid 432
(416 in the mature DBP) and one at amino acid 436 (420
in the mature DBP). The 1F allele encodes the sequence
of amino acids between 432 and 436 as DATPT, the
1S allele encodes the sequence EATPT. The Gc2 allele
encodes DATPK.
In addition to the polymorphisms, the level of glyco­
sylation further distinguishes the Gc1 variants from the
Gc2 variant. The threonine (T) in Gc1 binds N-acetyl­
galactosamine to which galactose and sialic acid bind
in tandem. The lysine (K) in comparable position in Gc2
does not bind these residues18,99. These differences affect
the conversion of DBP to DBP macrophage activation
Nature Reviews | Endocrinology
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factor (MAF), as only the glycosylated vari­ants can be
converted to DBP–MAF100. The different variants have
been shown to have different affinities for 25OHD101,
although the results and biological significances of dif-
ferences in affinities have not been consistent among
different laboratories102–105. That said, the affinities of
the DBP variants seem to be altered in different clini­
cal conditions. For example, in the third trimester of
pregnancy, directly measured free 25OHD tends to be
increased and free 1,25(OH)2D is substantially higher
in pregnant women than would be expected by meas-
uring total levels in comparison to non-pregnant
women106,107. This finding suggests that the affinity of
DBP for the vitamin D metabolites is decreased during
pregnancy. Similarly, directly measured free 25OHD
and 1,25(OH)2D tend to be higher in outpatients with
cirrhosis than in other groups107,108 despite the fact that
individuals with cirrhosis have reduced total vitamin D
metabolite concentrations108. This changed relationship
between total and free 25OHD levels can be appreciated
by the steeper but more variable slope in the correlation
between free 25OHD and total 25OHD in patients with
liver disease compared with that in healthy people. These
data indicate that there is a decreased affinity of DBP for
25OHD in these patients. A similar observation has been
made in patients in nursing homes109,110.
Allelic variation might also affect the affinity of
DBP for the vitamin D metabolites of DBP. Most assays
have shown a modest reduction in circulating levels
of 25OHD levels in individuals with the Gc2 variant of
DBP compared with those with the Gc1 variants111–114,
but the differences in directly measured free 25OHD lev-
els are nearly the same115. A genome-wide association
study reported in 2010 demonstrated that rs2282679, an
intronic polymorphism in the DBP gene, was likewise
associated with lower 25OHD and DBP levels40. On the
other hand differences in these alleles were not found
to contribute to a difference in fracture rate in a study
in 12,781 African American and white individuals116
or in patients with other calcaemic or cardiometabolic
diseases in the Canadian Multicentre Osteoporosis
study117. Nevertheless, a large number of chronic dis-
eases including type 1 and type 2 diabetes mellitus118–120,
osteoporosis121–123, chronic obstructive lung disease124,
endometriosis125, inflammatory bowel disease126, some
cancers127–130 and tuberculosis131 have been associated
with various DBP alleles.
Intracellular transport of vitamin D metabolites. The
primary role of DBP is to transport the vitamin D
metabolites between cells, but an intracellular transport
network also exists. What was originally called intracel-
lular DBP, now known as HSP70 (ref.132), promotes the
cellular uptake and distribution of vitamin D metabolites
within the cell23. Overexpression of this protein in cells
from Old World monkeys (MLA-144 cells) increased
the amount of 25OHD in these cells, promoted synthe-
sis of 1,25(OH)2D and increased the genomic actions
of 1,25(OH)2D as assessed by increased expression of
CYP24A1 (ref. 133) . Antisense constructs to HSP70
blocked these actions. The concept is that HSP70 func-
tions to transport 25OHD to the mitochondria where
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CYP27B1 (the 1-hydroxylase) converts the 25OHD to
1,25(OH)2D, which is then transported to the nucleus
where it binds the VDR for its genomic actions. HSP70
has higher affinity for 25OHD than DBP facilitating
25OHD transport across the cell membrane, but lower
affinity for 1,25(OH)2D than VDR, which facilitates
1,25(OH)2D offloading onto VDR in the nucleus133.
VDR
Tissue distribution. VDR, a member of the nuclear
receptor superfamily, was initially identified in the
intestine, the site which at the time was the best place to
study the action of vitamin D134. Although the highest
concentration of VDR is in tissues involved in the main-
tenance of calcium homeostasis (intestine, kidney and
bone), VDR has been found in many other tissues
and cells not involved in calcium homeostasis including
pancreas, skin, pituitary, breast, colon and prostate cancer
cells, and immune cells, suggesting that vitamin D affects
many cellular processes and a role for vitamin D beyond
regulation of calcium homeostasis135,136. The functions of
VDR in these other tissues and cells is a topic of ongoing
investigation, as the expression levels of VDR in different
cells are quite variable and can change with differentiation
of the cell135,137.
Side view Top view
RXRα LBD
RXRα LBD
VDR LBD VDR LBD
RXRα
DBD
VDR DBD VDR
RXRα DBD hinge
VDR
DBD
Fig. 3 | Vdr–rXrα vitamin d receptor binding site (dr3) model. The vitamin D receptor
(VDR)–retinoid X receptor-α (RXRα) heterodimer was created in the ‘open’ conformation,
and a partial cryo-electron microscopy model was used to guide the creation of the model.
The VDR–RXRα heterodimer forms an extended L-shaped organization with RXRα DNA
binding domain (DBD) occupying one half-site of DR3 (a synthetic oligonucleotide
representing an idealized direct repeat VDR binding site; shown in black in the figure)
at the 5′-end and VDR DBD occupying another at the 3′-end145. The VDR hinge domain,
a connection linker between the DBD and the ligand binding domain (LBD), plays a key
role in determining structural orientation. In this model, the VDR hinge domain adopts an
α-helical structure residing close to DR3 and makes extensive contacts with phosphate
and the backbone of DNA145. This is consistent with observations made with hydrogen–
deuterium exchange mass spectrometry showing that the VDR hinge undergoes different
extents of protection upon binding to various VDR binding sequences138. Unlike VDR ,
DNA binding does not perturb RXRα hinge dynamics as it forms a flexible linker that allows
enhanced adaptability of the heterodimer to diverse response elements138. This model
suggests that the hinge could possess intrinsic properties to orient the LBD dimer in a
precise way and provide a structural basis for crosstalk between LBDs and DBDs. VDR ,
RXRα and DNA are coloured in blue, grey and black, respectively. Image courtesy of
P. Griffin, Scripps Research Institute, USA.
VDR structure. The structure of VDR is shown in Fig. 3
(ref.138). The human VDR contains 427 amino acid res-
idues. The core domains of VDR are a highly variable
C terminal ligand-binding domain (LBD), a highly
conserved DNA binding domain (DBD) and a hinge
region connecting the DBD to the LBD. As indicated
by X-ray crystallography data, the DBD contains two
zinc fingers. Each zinc atom is held in a tetrahedral
configuration by four cysteine residues139. The LBD
is composed of 12 α-helices (H1–12)140. The terminal
H12 in the non-liganded VDR is open, but with ligand
binding H12 moves over the ligand and encloses it in
the binding pocket140. Along with H3 and H4, H12 (the
ligand-dependent activation function (AF2) corre-
sponds to H12) provides the interface for co-activator
binding140. The LBD of VDR is large and can accom-
modate a number of ligands including lithocholic acid
in addition to 1,25(OH)2D, although 1,25(OH)2D is the
preferred ligand as it has the highest affinity for VDR141.
Moreover, a model has been proposed in which the
VDR can accommodate two types of ligand configura-
tions — the genomic pocket for analogues with genomic
activity and the alternative pocket for analogues trigger-
ing rapid and non-genomic responses142. This model,
however, remains controversial.
In addition to X-ray crystallography data, which
provide a static view of VDR structure, complementary
data from NMR has provided insights into the confor-
mational changes that occur when VDR is bound to
different ligands. These conformational changes result
in differing biological activity143. Solution methodology
(such as X-ray scattering and cryo-electron microscopy,
as well as hydrogen–deuterium exchange mass spec-
trometry) has also yielded new information concerning
the structure and function of the VDR–RXR hetero­
dimer144–146. The VDR hinge region has been found to
be essential for stabilizing the VDR–RXR complex and
maintaining the integrity of the functional structure144,145.
Conformational dynamics indicate that the binding of
the heterodimer to DNA results in significant alterations
in the conformation of the LBD within regions that are
critical for interaction with co-activators146. Sequences
of the DNA response element and ligand structure dif-
ferentially affect the conformation of the heterodimer,
which can lead to selectivity of co-activator binding and
gene-specific effects of VDR138,146. Further studies using
solution structure methods for VDR complexes that are
difficult to crystallize will provide an increased under-
standing of the cooperative influence of 1,25(OH)2D,
DNA, VDR–RXR heterodimer and co-activator binding
on the mechanism of action of vitamin D.
Mutations of the VDR. When VDR is defective due to
mutations in VDR, the result is early onset of rickets
with resistance to 1,25(OH)2D. The resulting form of
rickets is termed hereditary vitamin D-resistant rickets
(HVDRR), which is also known as vitamin D-dependent
rickets type II. Patients have low serum calcium and
phosphate, high circulating levels of PTH and alkaline
phosphatase and very high levels of 1,25(OH)2D30. To
date, at least 49 different mutations in over 100 patients
have been described30,147. Mutations have been identified
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in the DBD and LBD, which inhibit VDR–RXR hetero­
dimerization30. These mutations result in bone defects
and alopecia areata30. Mutations in the LBD that alter
binding affinity for 1,25(OH)2D (for example, Arg­
274Leu, Arg274His and His305Gln) demonstrate the
important amino acids for contact points of 1,25(OH)2D.
The mutation in H12 (Glu420Lys) has been found not
to affect ligand binding, DNA binding or VDR–RXR
heterodimerization but prevents co-activator binding,
indicating the critical role of co-activator recruitment
by VDR to enable its function in preventing rickets30,147.
Mutations that disrupt ligand binding or co-activator
binding result in rickets but do not cause alopecia areata,
indicating that control of hair follicle cycling by VDR
is ligand-independent30,147. Mouse models of HVDRR
have been created that express human VDR mutants
in the LBD, in which 1,25(OH)2D binding is impaired in
the absence of endogenous mouse Vdr148–150. In these
VDR-mutant mice with impaired 1,25(OH)2D binding,
the alopecia areata but not the biochemical and skele-
tal aspects of the Vdr-null mice was rescued, confirm-
ing a role for unliganded VDR in hair follicle cycling.
Interestingly, skeletal defects in the mice expressing
ligand-binding-deficient VDR were more severe than
in the Vdr-null mice148,149. In addition, in a subset of
VDR target genes in the intestine (Trpv6, Cyp24a1 and
Slc30a10), gene downregulation in the Vdr-mutant
mice was stronger than in the Vdr-null mice, suggest-
ing suppression by VDR in the absence of 1,25(OH)2D3
(ref. 149) . Although VDR binding to DNA is mostly
ligand-dependent, these in vivo findings are notable
since they suggest that signalling pathways mediated
by unliganded VDR as well as ligand-dependent VDR
signalling can have a substantial role in calcium and
bone homeostasis.
Patients with HVDRR are treated with pharmacolog-
ical doses of vitamin D. Treatment with 25(OH)D3 or
1,25(OH)2D3 has been shown to benefit individuals with
mutations in the LBD that reduce binding affinity for
1,25(OH)2D147. However, if individuals fail to respond
to vitamin D or vitamin D metabolites, oral calcium
therapy or intravenous calcium infusion reverses all
aspects of HVDRR except alopecia areata151, which
indicates that the most important role of 1,25(OH)2D
and/or VDR in calcium homeostasis is regulation of
intestinal calcium absorption. As patients grow older,
the need for calcium supplementation lessens, suggest-
ing VDR-independent pathways for intestinal calcium
absorption in patients with HVDRR later in life147,152.
Regulation of VDR. The multiple actions of
1,25(OH)2D that are mediated by VDR involve bind-
ing of 1,25(OH)2D and VDR–RXR to specific DNA
sequences in the regulatory regions of target genes
and the recruitment of co-regulatory complexes,
which results in the modulation of gene expression153.
Co-regulatory factors involved in VDR-mediated tran-
scription include factors with histone acetylase activity,
including steroid receptor co-activator 1 (SRC1), SRC2
and SRC3, and CREB-binding protein p300. Additional
co-regulatory factors include SWI–SNF ATP-dependent
chromatin-remodelling complex, methyltransferases
Nature Reviews | Endocrinology
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and the mediator (MED) complex, which functions to
recruit RNA polymerase II136,153,154. VDR binding sites
are associated with sites for other transcription factors
such as C-EBPα, C-EBPβ, RUNX2 and PU.1, which can
cooperate with VDR and VDR co-regulators to influence
1,25(OH)2D responses in target cells155–157. Genome-wide
studies have shown that VDR binding sites are located
within introns, in intergenic regions and many kilobases
upstream or downstream of regulated genes as well as in
promoters adjacent to transcription start sites153.
Consistent with the ability of VDR to recruit co-
regulatory complexes that regulate chromatin structure,
evidence provided over the past several years shows that
levels of acetylation at H4K5, H3K9 and H3K27 define
sites of action of 1,25(OH)2D and occur within enhanc-
ers that regulate 1,25(OH)2D target genes158. Specific
enhancer deletions in vivo have provided an important
advance in the determination of tissue-specific determi­
nants of VDR actions. These studies using loss-of-
function enhancer deletion studies in mice have identi­fied
a kidney-specific multicomponent control module distal
to Cyp27b1 that controls basal and hormone-regulated
expression of Cyp27b1 in the kidney, but that is not active
in other cells such as immune cells which use another
module conferring cytokine regulation. These data
provide an insight, for the first time, into cell-specific
mecha­nisms involved in the regulation of CYP27B1 that
had remained undefined27.
Studies involving genome-scale definition of VDR
action have also shown the translational potential of
VDR as a drug target. The VDR ligand calcipotriol
(a less-hypercalcaemic analogue of 1,25(OH)2D) was
found to suppress profibrotic gene expression in liver
stellate cells through antagonism of SMAD3 recruit-
ment, thus defining a role for VDR in the regulation
of liver fibrogenesis159. VDR was also found to act as a
master transcriptional repressor of the pancreatic stel-
lar cell activation state, which suggests that reprogram-
ming by vitamin D could be an adjunct in pancreatic
ductal adenocarcinoma therapy160. In addition, VDR
was found to have a role in β-cell survival by reversing
inflammation-driven transcriptional changes161. VDR
ligand was shown to promote VDR association with the
PBAF chromatin remodelling complex, which enhanced
chromatin accessibility at vitamin D response elements
resulting in inhibition of inflammatory response genes161.
These findings indicate that VDR transcriptional
mechanisms are involved in β-cell survival. Further stud-
ies involving genome-scale definition will reveal addi-
tional novel mechanisms of 1,25(OH)2D action that could
result in new approaches not only to sustain calcium bal-
ance, but also to enhance the suggested antitumour and
anti-inflammatory actions of 1,25(OH)2D (Box 2).
The role of vitamin D in skin
The epidermis and hair follicle are models of vitamin D
signalling that demonstrate the role of vitamin D beyond
that of regulating bone mineralization and intestinal
calcium transport. Aside from producing vitamin D,
epidermal cells (keratinocytes) convert the vitamin D to
its biologically active metabolite 1,25(OH)2D162. More­
over, keratinocytes contain VDR163,164, and respond to
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1,25(OH)2D with changes in proliferation and differenti-
ation31,165,166. Vitamin D signalling in keratinocytes works
in partnership with calcium, acting in part through
the calcium-sensing receptor (CaSR) and β-catenin1.
Deletion of VDR and/or CaSR in epidermal keratino-
cytes (epiVdr-null and/or epiCasr-null) disrupts epidermal
differentiation167,168, impairs wound healing169,170 and pre-
disposes to cancer171. Although VDR is generally thought
to function in combination with its ligand 1,25(OH)2D,
hair follicle cycling is one notable exception. Vdr-null
animals develop loss of hair follicle cycling172,173, whereas
those lacking Cyp27b1 or Casr, or those placed on a
vitamin D-deficient diet, do not1. On the other hand,
β-catenin signalling is an important partner with VDR
in regulating hair follicle cycling26,174 and stem cell acti-
vation as during the response to wounding169. Within
the epidermis, β-catenin has a dual role in enabling
proliferation as a transcription factor but facilitating
differentiation as a key component of the membrane
E-cadherin–catenin complex (Box 3).
Regulation of keratinocyte differentiation. The epider-
mis is an excellent example of differentiation (Fig. 4). The
basal layer (stratum basale), which rests on the basal
lamina (the layer that separates the dermis and the epi-
dermis), is where the interfollicular epidermal stem cells
reside175. These stem cells give rise to transient amplify-
ing cells that begin the differentiation process as they
move upward into the next layer, the stratum spinosum1.
Cells in this layer produce keratins K1 and K10 (kera­
tins in the basal layer are K5 and K14)176. Moreover, one
of the important components of the cornified envelope,
involucrin177, is produced in the stratum spinosum along
with the enzyme transglutaminase K, which is respon-
sible for the ε-(γ-glutamyl)lysine crosslinking of involu-
crin and other substrates (for example, loricrin) into the
insoluble cornified envelope178.
Above the stratum spinosum is the stratum granu­
losum, which as its name implies is characterized by
electron-dense keratohyalin granules179. The larger gran-
ules contain profilaggrin180. Profilaggrin is the precursor of
filaggrin, which is thought to facilitate the aggregation
of keratin filaments181, but when hydrolysed is thought
also to provide a moisturizing factor for the skin182. The
smaller granules contain loricrin, which like involucrin
is an important component of the cornified envelope183.
Box 2 | Key concepts in the regulation of tissue-specific actions of Vdr
• Conformational dynamics have shown that the vitamin D receptor (VDR)–retinoid X
receptor (RXR) heterodimer adopts different conformations depending on sequences
in the DNA response element, ligand structure and co-regulatory protein binding138.
• Genome-wide studies have noted that actions of 1,25(OH)2D involve regulation of
gene activity at a range of locations. VDR binding may be many kilobases upstream body formation, in a similar way to Vdr deletion193, and
or downstream of regulated genes154.
• Sites of action of 1,25(OH)2D generally occur within enhancers that regulate
1,25(OH)2D target genes154.
• Loss of function enhancer deletion studies in mice have identified a multicomponent
control module that controls basal and hormone regulated expression of Cyp27b1.
• Genome-scale definition of VDR action has noted the translational potential of from a proliferative state to one of differentiation1. This
VDR as a drug target in liver fibrogenesis, pancreatic ductal adenocarcinoma and
type 2 diabetes154.
Lamellar bodies, which contain both long-chain fatty
acids and antimicrobial peptides from the stratum gran­
ulosum, line the membrane separating the stratum
granulosum and the stratum corneum184. The lamellar
bodies are positioned to offload their contents into the
extracellular space where the lipids and antimicrobial
peptides contribute to the permeability barrier and
defence against microorganisms185,186.
1,25(OH)2D with its receptor VDR regulate all steps
in the differentiation process. For example, 1,25(OH)2D
induces keratin 1, involucrin and transglutaminase
activity in the stratum spinosum, and filaggrin, loricrin,
antimicrobial peptide and long-chain fatty acid synthesis
and cornified envelope formation in the stratum granu-
losum165,166,187–190. In addition to its role in epidermal dif-
ferentiation, 1,25(OH)2D promotes the innate immune
function of keratinocytes by inducing the Toll-like
receptor 2 (TLR2) and its co-receptor CD14 (ref.191).
The induction of TLR2 and CD14 in turn has a feedback
element in that activation of TLR2 and CD14 induces
CYP27B1, which increases production of 1,25(OH)2D
that then induces cathelicidin, a potent antimicrobial
peptide191,192. Deletion of either Vdr or Cyp27b1 in mice
results in defective epidermal differentiation, a defective
permeability barrier193, and a defective response of the
innate immune system to infection and wounding191,194.
The process of epidermal differentiation and its
regulation by 1,25(OH)2D and VDR is sequential in
that different genes and pathways are induced in the
keratinocytes at their different states of differentia-
tion. VDR and CYP27B1 are expressed throughout
the epidermis, although their highest expression is in the
stratum basale195,196. As noted previously, the transcrip-
tional activity of VDR is tightly regulated by a number
of co-regulators that can show cell specificity136,153. The
major co-activator complexes in the epidermis regulat-
ing VDR function are MED (formerly called DRIP or
VDR interacting proteins) of which MED1 is the major
VDR-binding component, and the SRC complex, of
which SRC2 and SRC3 are the major VDR-binding com-
ponents197. The functions of VDR in undifferentiated
keratinocytes are controlled primarily by MED, whereas
the actions of VDR in the more differentiated cells are
regulated by SRC2 and SRC3 (refs197,198). MED1 and
SRC3 are reciprocally expressed in the epidermis, with
MED1 primarily found in basal keratinocytes and SRC3
in the more differentiated layers, that is, the stratum
granulosum197. Deletion of Med1 but not Src3 in mice
results in increased keratinocyte proliferation198 and
a profound disruption of hair follicle cycling199, and has a
greater effect on K1, K10 and involucrin expression than
does the deletion of Src. On the other hand, deletion of
Src3 reduces glucosylceramide production and lamellar
prevents 1,25(OH)2D-induced cathelicidin expression200.
As alluded to earlier, calcium partners with
1,25(OH)2D and/or VDR in their effects on prolifera-
tion and differentiation. Increases in the extracellular
calcium concentration above 0.1 mM change the cells
change in state includes the development of intercellu-
lar contacts, the formation of the E-cadherin–catenin
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Box 3 | The skin as a model for the study of vitamin d
• The skin is the major source of vitamin D as it contains a large store of
7-dehydrocholesterol for conversion to vitamin D upon UVB radiation exposure6.
• All the enzymes required for vitamin D metabolism are present in the keratinocytes tion218,219, a role that may be important in the regulation
of the epidermis and hair follicle. Regulation of at least CYP27B1, but most likely
CYP24A1 as well, differs from that in the kidney1.
• Keratinocytes express vitamin D receptor (VDR), and so respond to its ligand
1,25(OH)2D with changes in proliferation, differentiation, barrier formation and
immune function. However, at different stages in the differentiation process different
co-regulators of VDR are required1,197.
• Hair follicle cycling is the best example known of a VDR-regulated process not
requiring 1,25(OH)2D173.
• Lack of VDR in the epidermis and hair follicle predisposes to deficient wound healing
and cancer169,255.
complex containing phosphoinositide 3-kinase (PI3K),
α-catenin and β-catenin, and phosphatidyl inositol 4-
phosphate 5-kinase 1α (PIP5K1α)1. All of these compo-
nents have critical roles in the differentiation process.
For example, PI3K and PIP5K1α are involved in the for-
mation of PIP3 in the membrane, which among other
functions, activates phospholipase Cγ1 (PLCγ1), a key
enzyme in maintaining the increased intracellular cal-
cium required for differentiation201. In addition to these
functions, the calcium switch leads to the membrane
localization of the CaSR, and SRC kinases, which also
have important roles in the calcium-induced differen-
tiation process by helping form the E-cadherin–catenin
complex202–207. α-Catenin links the E-cadherin–catenin
complex to the actin cytoskeleton enabling cell migra-
tion. CaSR is essential for the keratinocyte response to
calcium204,208. Deletion of Casr from keratinocytes blocks
their response to extracellular calcium by reducing their
stores of calcium and preventing the formation of the
E-caderin–catenin complex204,209. VDR and/or CYP27B1
and CaSR are mutually required for each other’s full
expression168,210, and participate in the joint regulation
of the expression of a number of genes by calcium and
1,25(OH)2D204,211–213.
The precise interaction between VDR and β-catenin
is dependent on which cells are involved. Both VDR and
β-catenin are required for hair follicle cycling, but their
interactions in the interfollicular epidermis are more
complex214. As noted previously, although 1,25(OH)2D is
not required for VDR regulation of hair follicle cycling,
β-catenin is. Deletion of Ctnnb1 (encoding β-catenin)
from keratinocytes in mice results in a nearly identical
phenotype as Vdr deletion with loss of hair follicle cycling
combined with sebocyte hypertrophy, dermal cysts lined
with epithelia expressing epidermal markers, and epi-
dermal hyperplasia172,215. These results indicate different
roles for VDR and β-catenin in the stem cells of the hair
follicle, sebaceous gland and interfollicular epidermis216.
β-Catenin interacts with the AF2 domain of VDR in a
ligand-dependent manner (like other co-activators) but
lymphoid enhancer binding factor 1 (LEF1; an impor-
tant transcription factor in β-catenin signalling) binds to
the N terminus of VDR in a ligand-independent man-
ner217 possibly essential for hair follicle cycling by these
transcription factors. The significance of these inter­
actions are discussed further later in the Review where
Nature Reviews | Endocrinology
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we consider the roles of vitamin D signalling in hair
follicle cycling, skin cancer and wound healing.
The ΔNp63α and other mutant forms of p53 have
been implicated in vitamin D regulation of prolifera-
by VDR in cancer. Similarly, VDR has been implicated
as a master regulator of the MYC/MXD1 network220,
playing a possible role in keratinocyte proliferation and
cancer. The E3 ubiquitin ligase MDM2 has been found
to bind to and decrease the function of VDR, potentially
contributing to uncontrolled proliferation and cancer221.
Further investigation of their roles in vitamin D signal-
ling in the skin is required to fully understand their level
of participation.
Hair follicle cycling. The hair follicle cycle is composed
of three distinct phases. The phase of hair follicle growth
is called anagen. Anagen is initiated following contact
between the hair follicle and the dermal papilla, a spe-
cialized collection of mesenchymal cells in the dermis222.
At a given point following the growth phase, the follicle
regresses — this is called catagen222. The trigger for cata­
gen, however, is not clear. Following catagen, the hair
follicle enters a resting phase called telogen222. Following
the developmental cycle, which leads to the initial coat
of hair, the hair follicle undergoes repetitive cycling
until senescence222. Although signals from the dermal
papilla are clearly involved in the reinitiation of anagen,
the length of telogen and the timing of the new cycle
are not well understood and vary between the sexes
and different parts of the body222. Only the proximal or
dermal portion of the hair follicle cycles; the distal
or epidermal portion does not222.
VDR regulation of the hair follicle cycle begins only
after the developmental hair follicle cycle is completed,
so that the initial coat of hair initiated during embryo­
genesis proceeds normally in Vdr-null mice172. Hair fol-
licle cycling is dependent on stem cells in the bulge, a
specialized region of the outer root sheath (ORS) below
the sebaceous gland223. These cells have a higher level
of VDR expression than other cells in the hair follicle.
The ORS is a direct extension of the stratum basale, and
separates the hair follicle from the surrounding connec-
tive tissue sheath224. The ORS is home to other stem cell
niches, including cells supporting the sebaceous gland
and the epidermis itself 225. The stem cells that support
the sebaceous gland and the epidermis are crucial in the
epidermal response to wounding225.
A marked characteristic of many patients with muta-
tions in VDR226,227 is alopecia, as is the case in Vdr-null
mice173. The block in hair follicle cycling occurs at the
point of reinitiation of anagen following the develop-
ment cycle228. This loss of hair follicle cycling is specific
for deletion of Vdr in the keratinocyte229, as demon-
strated in a conditional mouse knockout model that
had none of the metabolic changes seen in the global
Vdr-knockout models.
There are, however, mutations in other signal-
ling mechanisms that are phenocopies of the alope-
cia in Vdr-null mice (without the metabolic changes).
These mutations provide insight into the role of VDR
in its regulation of hair follicle cycling. Mutations in
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Lamellar body
Keratohyalin
granule
Stratum corneum
Cornified envelope
SRC3
Stratum granulosum
Profilaggrin and/or
loricrin, cathelicidin
and lipid production
Stratum spinosum
INV and/or TG or
K1 and/or K10
MED1
Stratum basale
• K5 and/or K14
• VDR, CYP27B1
• Stem cells
Fig. 4 | The four layers of the epidermis, their distinct functions and location of
major Vdr co-regulators. The epidermis is composed of four layers. The stem cells are
found in the stratum basale, where the keratins K5 and K14 are expressed. Here are also
the highest concentrations of vitamin D receptor (VDR) and CYP27B1. As the progeny
of the stem cells move up from the basal layer they become part of the stratum spinosum.
Here, an important component of the cornified envelope, involucrin (INV), is produced
along with transglutaminase (TG) that crosslinks INV and other proteins into the insoluble
cornified envelope. The main keratins expressed in the stratum spinosum are K5 and K10.
The stratum granulosum lies just under the stratum corneum and expresses a number of
proteins such as loricrin (in the keratohyalin vesicles) that also form part of the cornified
envelope, and profilaggrin, that originally was thought to bundle the keratins but also
provides peptides for incorporation into the lipid envelope lying beneath the cornified
envelope. The stratum granulosum also expresses the enzymes for processing long-chain
fatty acids that are incorporated into lamellar bodies along with defensins such as
cathelicidin that are secreted into the lipid envelope and cornified envelope to both
‘waterproof’ the cornified envelope and provide defence against invading organisms.
1,25(OH)2D/VDR is a key regulator of proliferation of the stem cells and their subsequent
differentiation as the cell moves from the stratum basale to the stratum corneum. This
change in 1,25(OH)2D/VDR-regulated gene expression during the differentiation process
is regulated by a shift in co-regulators of which the mediator complex is critical for
the early stages and steroid receptor co-activator 3 (SRC3) for the later stages of the
differentiation process. MED1, mediator complex subunit 1.
Hairless (Hr) in both mice230 and humans231,232, as well
as transcriptionally inactivating Ctnnb1 mutations in
mice222,233, are two such examples. Hairless (Hr) binds
to VDR and in epidermal keratinocytes suppresses its
1,25(OH)2D-stimulated transcriptional activity234–236. The
role of Hr in ligand-independent VDR regulation of hair
follicle cycling is less clear. β-Catenin can enhance VDR
signalling especially in the hair follicle, whereas VDR can
inhibit β-catenin transcriptional activity in the epider-
mis174,214,237. In mice with these different mutations that
cause aborted hair follicle cycling, the pathological pro-
cess begins during catagen at the end of the developmen-
tal cycle, the dermal papilla dissociates from the hair
follicle, and anagen is not reinitiated172,230,238. Signalling
via the Hedgehog and β-catenin pathways is required for
these mesenchymal and/or epithelial interactions, and
they interact with each other and with VDR to enable hair
follicle cycling239. Hedgehog signalling is mediated by the
transcription factors GLI1 and GLI2, whereas β-catenin
signalling involves LEF1. The expression of GLI1 and
GLI2 is reduced during the latter stages of catagen in
both the Hr- and VDR-null hair follicle171. On the other
hand, expression of Hedgehog signalling components
is increased in the interfollicular epidermis and utricles
of the Vdr-null mouse240, but their expression is blocked
by 1,25(OH)2D229. These observations emphasize the
differences in signalling between the cycling portion of
the hair follicle and that in the non-cycling portion
of the hair follicle and epidermis. As noted previously, in
the Vdr-null mouse as well as in Hr-mutant and Ctnnb1-
mutant animals222,233,238,241, large lipid-filled dermal cysts
develop that contain markers of the differentiated inter-
follicular epidermis along with hypertrophy of the seba-
ceous glands215,238,242. These findings suggest that changes
in cell fate of the stem cells within these niches differ
from those in the bulge.
Alopecia areata, as opposed to the alopecia totalis
found in Vdr-null mice (and humans), is often accom-
panied by vitamin D deficiency, and data show that it can
be improved by vitamin D supplementation243,244. This
condition, however, is thought to be due to immune
destruction of the hair follicle, with 1,25(OH)2D serv-
ing to suppress this process similar to the mechanism
in other inflammatory diseases of the skin such as pso-
riasis245. A transient disruption of hair follicle cycling
was observed in S100g-knockout mice raised to wean-
ing by mothers on a vitamin D-deficient low-calcium
diet. Normal hair follicle cycling was restored after
weaning246. The explanation for this phenomenon is not
clear, but might point to a role of calcium in hair folli-
cle cycling not observed in mice lacking Casr in their
keratinocytes. Med1 deletion leads to an acceleration of
hair follicle cycling during the initial cycles, opposite to
the effect of Vdr deletion, but hair follicle differentiation
is disrupted and, as is the case with Vdr deletion, Med1
deletion results in a gradual loss of bulge stem cells199.
Epidermal cancer development
Skin cancer is the most common form of cancer247. Most
skin cancers are basal cell carcinomas (BCC; 80%) and
squamous cell carcinomas (SCC; 16%), with the more
deadly melanomas comprising about 4%. In individuals
with these cancers, circulating levels of 25OHD are not
deficient and might even be increased248–250. BCC and
SCC are currently referred to as keratinocyte carcinomas.
Moreover, these cancer cells can produce 1,25(OH)2D
and 24,25(OH)2D, with rates of production that can be
comparable to those of normal keratinocytes in some
cases188. However, when tested in vitro, keratinocyte car-
cinoma cells lack a normal response to either calcium or
1,25(OH)2D188,250,251, suggesting a mechanism other than
loss of VDR or CYP27B1 that is disrupted. As discussed
previously, MED and SRC co-activator complexes regu­
late the sequential actions of VDR and/or 1,25(OH)2D
on keratinocyte proliferation and/or differentiation with
the shift from MED to SRC being required for terminal
differentiation of the cell. The keratinocyte carcinoma
cell lines we have studied overexpress MED1, with little
or no expression of SRC3, which suggests that this blocks
the ability of VDR to promote the differentiation process
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by blocking access to SRC3 required for induction of the
more differentiated functions of the keratinocyte197.
A second explanation stems from the reduction in
formation of the E-cadherin–catenin complex in kera­
tinocyte carcinomas. As noted previously, formation
of the E-cadherin–catenin complex is a key step in
keratinocyte differentiation. Moreover, failure to form
this complex, which would maintain β-catenin in the
membrane, enables more β-catenin to translocate to
the nucleus, thereby promoting proliferation and
bloc­king differentiation. Additionally, there is a shift
in the mode of activation of PLCγ1 from PIP3 (via the
E-cadherin–catenin complex), a mode of activation that
promotes the ability of PLCγ1 to maintain elevated intra-
cellular calcium levels required for differentiation, to its
activation by growth factors such as epidermal growth
factor, shifting its role to promoting proliferation252.
Although these mechanisms may help explain the
lack of response to 1,25(OH)2D in cancers that express
the VDR, lack of VDR predisposes to the development of
keratinocyte carcinomas in otherwise normal keratino­
cytes. Mice null for Vdr develop tumours (papillomas
and keratinocyte carcinomas) at a much higher rate than
controls when exposed to carcinogens (7,12-dimethyl­
benzanthracene)253 or UVB radiation229,254. Moreover,
mice lacking both Vdr and Casr were found to develop
tumours spontaneously255. That said, linking vitamin D
signalling to keratinocyte carcinomas in humans has
been difficult, and the results of epidemiological studies
have been mixed256,257. That said, several mechanisms,
not mutually exclusive, underlying this predisposition to
keratinocyte carcinoma formation at least in mice have
been proposed258, and are covered briefly here.
Regulation of DNA damage response. UVB radiation
(280–320 nm) induces the formation of cyclobutane
pyrimidine dimers and pyrimidine primidone photo-
products in the epidermis that if not repaired result in C
to T or CC to TT mutations — which are the UVB radi-
ation signature lesions259. UVA radiation (320–400 nm)
damages DNA primarily by oxidative processes (such as
8-hydroxy-2′-deoxyguanosine production). Reversing
these lesions is the role of DNA damage response oper-
ating primarily by nucleotide excision repair (NER)260,261.
This process is defective in the epidermis of Vdr-null
mice 229,262. In normal mice, topical application of
1,25(OH)2D seems to be effective in preventing these
lesions and/or expediting their repair263. 1,25(OH)2D
induces two genes important for NER — XPC and DDB2
(ref.264) — but in both cases VDR is required.
Regulation of the Hedgehog pathway. Overexpression of
Hedgehog signalling due to mutations in PTCH1 or other
genes of the Hedgehog pathway is a well-documented
cause of nearly all BCC265–267. The Hedgehog pathway
leads to the activation of the GLI family of transcription
factors, which in turn increase the expression of each
other as well as PTCH1, and a variety of antiapoptotic
proproliferative factors while suppressing expression
of proteins associated with keratinocyte differentia-
tion268–270. This pathway is overexpressed in the epider-
mis and epidermal portion (utricles) of the hair follicle of
Nature Reviews | Endocrinology
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adult Vdr-null mice240, unlike the cycling part of the hair
follicle in which the Hedgehog pathway is suppressed in
Vdr-null mice. 1,25(OH)2D and/or VDR suppresses the
expression of components of the Hedgehog pathway240,
presumably through the VDR response elements found
in their promoters174,217. Surprisingly, vitamin D by itself
seems to directly inhibit Smo, a key member of the
Hedgehog pathway7.
Regulation of β-catenin signalling. Although the
tumours (pilomatricomas or trichofolliculomas) that
result from overexpression or activating mutations in the
β-catenin pathway are not malignant271, when combined
with the deletion of VDR, BCC develops174. This finding
suggests that VDR restrains the potentially oncogenic
actions of β-catenin via the mechanisms discussed above
in the section describing the interactions between VDR
and β-catenin in keratinocytes. Moreover, in a number
of genes response elements for both β-catenin and VDR
have been found in close proximity174, which suggests
dual regulation of these genes that in the absence of VDR
might be oncogenic.
Regulation of long non-coding RNAs. Long non-coding
RNAs (lncRNAs) are endogenous cellular RNAs larger
than 200 bases that account for 80% of the transcriptome272.
They regulate many aspects of cellular physiology273,
including cancer development. We found that Vdr dele-
tion in cultured keratinocytes and in the epidermis in vivo
led to increased expression of oncogenic lncRNAs with
decreased expression of tumour suppressor lncRNAs274.
Regulation of the hyaluronan–CD44 pathway.
Hyaluronan, a major glycosaminoglycan within the
extracellular matrix, binds to CD44 (ref.275), a function-
ally important membrane receptor found in most cells
including keratinocytes. CD44 is encoded by 19 exons,
but is expressed in numerous forms due to alternative
splicing, in particular of exons 6–14 (ref.276). Differen­
tiated keratinocytes express a more complete form of
CD44 (known as epican), whereas undifferentiated
keratinocytes, mouse skin after chronic UVB irradiation,
and SCCs express a variety of shorter CD44s that signal
differently277.
Hyaluronic acid is synthesized by different hyal­
uronic acid synthases and is degraded by different hyal­
uronidases. UVB radiation alters both hyaluronic acid
synthesis and its degradation 278. Large hyaluronic
acid predominates in normal mouse skin, whereas small
hyaluronic acid predominates in cancer tissue279. Large
hyaluronic acid promotes transcriptional activation and
differentiation, whereas small hyaluronic acid induces
the expression of pro-inflammatory cytokines and/or
chemokines as well as cell proliferation and migration280.
We have proposed that the large hyaluronic acid acting
on epican promotes keratinocyte differentiation and
DNA repair through RAC–PKNγ and p38 MAPK sig-
nalling, whereas small hyaluronic acid acting on shorter
CD44 variants promotes proliferation and inflamma-
tion through RhoA–ROK-dependent NF-κB–STAT3
signalling281. 1,25(OH)2D blocks small hyaluronic acid–
CD44-mediated RhoA–ROK activation and NF-κB–p65
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signalling as well as inflammatory gene expression and
proliferation in transformed keratinocytes and SCC281.
Regulation of VDR levels and function. Tumours that
are unresponsive to vitamin D have typically lost their
ability to produce 1,25(OH)2D (caused by decreased lev-
els of CYP27B1)282,283. Furthermore, these tumours may
have an increase in the metabolism of 1,25(OH)2D via
upregulation of CYP24A1 (ref.284), lost VDR transcrip-
tional activity through post-translational alterations
in RXR285, or a decrease in VDR expression284 — the
last of which might be secondary to increased activity
in inhibitors of VDR expression such as SNAIL286 and
SLUG287, increased methylation of the VDR promoter288,
or increased expression of miRNA125b (an inhibitor of
VDR expression)289.
p53 cooperates with VDR in its antitumour func-
tions290. The nuclear levels of p53 are increased by
1,25(OH)2D by non-genomic mechanisms291. Never­
theless, p53 mutations in tumours can also contribute
to resistance to vitamin D in tumours. The E3 ubiqui-
tin ligase and transcriptional regulator MDM2 is also
regulated by 1,25(OH)2D292, and unlike p53 MDM2
inhibits VDR and transcriptional activity293, as well as
p53. Moreover, not all actions of VDR are genomic.
As noted previously, the 1,25(OH)2D stimulation of p53
is non-genomic. Analogues of 1,25(OH)2D that do not
have genomic activity protect the skin from the UVB
radiation-induced formation of cyclobutane pyrimidine
dimers294. These rapid effects of 1,25(OH)2D and its
non-genomic analogues are mediated by either or both
VDR acting in the membrane and an unrelated receptor
known initially as MARRS, but also known as ERp57,
GRp58, ERp60 or PSIA3.
The role of vitamin D in skin cancer is likely to be
multifaceted295. Given the definitive role that Hedgehog
signalling has in BCC, the potential role that vitamin D
plays in suppressing this pathway is a compelling ration-
ale for its use as a therapeutic agent. Moreover, the
potential role of vitamin D in promoting DNA damage
repair might provide a rationale for its use in the preven-
tion of UVB radiation-induced skin cancer (both SCC
and BCC). That said, at this stage of our knowledge it is
premature to judge which mechanisms are likely to be
most important in the antitumour actions of vitamin D.
Epidermal wound repair
Wound healing is a multistage process. Inflammation
and activation of the innate immune system are
important early responses. Keratinocytes null for Vdr,
Cyp27b1 or Casr have blunted inflammatory and innate
immune responses to wounding168,191. Wounding also
leads to a rapid increase in intracellular calcium170 and
β-catenin signalling169. In response to the cytokines
and growth factors expressed during this initial stage,
stem cells in the three niches of the hair follicle and
interfollicular epidermis are activated and proliferate,
and their progeny migrate to re-epithelialize the wound
over the matrix re-formed by dermal fibroblasts296.
Although stem cells from all three niches contribute
to re-epithelialization, the majority of cells perma-
nently repopulating the epidermis originate from the
interfollicular epidermis and infundibulum of the hair
follicle rather than the bulge.
Oda and colleagues 169,297 observed delayed re-
epithelialization of wounds in keratinocyte-specific
Vdr-null mice on a low calcium diet or mice with com-
bined Vdr and/or Casr deletion. Similar results were
seen in mice lacking Casr in their keratinocytes170.
These results were associated with reduced numbers
of stem cells, a reduction in their activation and migra-
tion over the newly formed matrix, and a subsequent
failure of these cells to differentiate to re-form the epi-
dermis. Others have noted a reduction in the number
and/or activation of bulge stem cells in global Vdr-null
mice237,298, although not in the context of wounding.
In the studies conducted by Oda and colleagues169,297,
β-catenin signalling was reduced as was the formation
of the E-cadherin–catenin complex in the migrating cells
with failure of Vdr-null keratinocytes to participate in
the re-formation of the epidermis. Stem cell activation
requires β-catenin signalling216 and, probably, calcium
signalling, whereas migration and/or differentiation
requires the E-cadherin–catenin complex201. In studies
of global Vdr-null mice on a high-calcium rescue diet,
Demay and colleagues299 observed that TGF signalling
in the dermis was reduced, associated with a reduction in
macrophage recruitment and granulation tissue forma-
tion. Thus vitamin D and calcium signalling have major
roles in all aspects of wound healing.
Conclusion
In this Review we provide an up-to-date discussion of
vitamin D metabolism, with a focus on lessons that can
be learned by studying mutations in the genes encod-
ing enzymes that enable vitamin D production, acti-
vation and catabolism. We also discuss the means by
which vitamin D and its metabolites are transported
from sites of production to sites of metabolism and
biological action. This portion of the Review focuses
on DBP with its numerous polymorphisms that bear on
its function. Within the cell HSP70 acts as a shuttle to
first help unload the vitamin D metabolites (25OHD in
particular) from DBP and shuttle the metabolite to the
mitochondria for further processing, then to transport
the active metabolite 1,25(OH)2D into the nucleus for
binding to VDR133.
It is VDR that has the critical biological role in medi-
ating the actions of 1,25(OH)2D. These actions, how-
ever, are cell- and tissue-specific, indicating that these
actions of 1,25(OH)2D/VDR are tightly regulated by a
number of factors including co-regulators and epige-
netic modifications of the DNA that dictate which genes
can be induced (or suppressed).
Following a description of newer insights into vita-
min D metabolism, transport and molecular mecha-
nisms of action, we then discuss an excellent model by
which these actions can be illustrated in a non-skeletal
tissue — the skin, in particular the epidermis and hair
follicle. These are regenerative organs in which the key
cell, the keratinocyte, has all the apparatus needed to
produce, metabo­lize and respond to vitamin D and
its metabolites1. We examine the important roles of
1,25(OH)2D and VDR in epidermal differentiation, the
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mechanism by which VDR without its ligand controls
hair follicle cycling, and the key co-regulators that dif-
ferentially regulate VDR action in the hair follicle and
epidermis. Although it is tempting to speculate that vita-
min D produced locally in the skin has a greater effect
on these functions than vitamin D ingested in food, this
1. Bikle, D. D. Vitamin D and the skin: physiology and
pathophysiology. Rev. Endocr. Metab. Disord. 13,
3–19 (2012).
2. Jäpelt, R. B. & Jakob sen, J. Vitamin D in plants:
a review of occurrence, analysis, and biosynthesis.
Front. Plant. Sci. 4, 136 (2013).
3. Tripkovic, L. et al. Comparison of vitamin D2 and
vitamin D3 supplementation in raising serum
25-hydroxyvitamin D status: a systematic review and
meta-analysis. Am. J. Clin. Nutr. 95, 1357–1364
(2012).
4. Tian, X. Q. & Holick, M. F. Catalyzed thermal
isomerization between previtamin D3 and vitamin D3
via β-cyclodextrin complexation. J. Biol. Chem. 270,
8706–8711 (1995).
5. Passeron, T. et al. Sunscreen photoprotection and
vitamin D status. Br. J. Dermatol. 181, 916–931
(2019).
6. Prabhu, A. V., Luu, W., Li, D., Sharpe, L. J. &
Brown, A. J. DHCR7: a vital enzyme switch between
cholesterol and vitamin D production. Prog. Lipid Res.
64, 138–151 (2016).
7. Bijlsma, M. F. et al. Repression of smoothened
by patched-dependent (pro-)vitamin D3 secretion.
PLOS Biol. 4, e232 (2006).
8. Bhattacharyya, M. H. & DeLuca, H. F. Subcellular
location of rat liver calciferol-25-hydroxylase.
Arch. Biochem. Biophys. 160, 58–62 (1974).
9. Horst, R. L., Littledike, E. T., Riley, J. L. & Napoli, J. L.
Quantitation of vitamin D and its metabolites and
their plasma concentrations in five species of animals.
Anal. Biochem. 116, 189–203 (1981).
10. Zhu, J. G., Ochalek, J. T., Kaufmann, M., Jones, G. &
Deluca, H. F. CYP2R1 is a major, but not exclusive,
contributor to 25-hydroxyvitamin D production
in vivo. Proc. Natl Acad. Sci. USA 110, 15650–15655
(2013).
11. Cheng, J. B., Levine, M. A., Bell, N. H.,
Mangelsdorf, D. J. & Russell, D. W. Genetic evidence
that the human CYP2R1 enzyme is a key vitamin D
25-hydroxylase. Proc. Natl Acad. Sci. USA 101,
7711–7715 (2004).
12. Bikle, D. D., Patzek, S. & Wang, Y. Physiologic and
pathophysiologic roles of extra renal CYP27b1: case
report and review. Bone Rep. 8, 255–267 (2018).
13. Thacher, T. D., Fischer, P. R., Singh, R. J., Roizen, J. &
Levine, M. A. CYP2R1 mutations impair generation of
25-hydroxyvitamin D and cause an atypical form of
vitamin D deficiency. J. Clin. Endocrinol. Metab. 100,
E1005–E1013 (2015).
14. Fu, G. K. et al. Cloning of human 25-hydroxyvitamin
D-1α-hydroxylase and mutations causing vitamin
D-dependent rickets type 1. Mol. Endocrinol. 11,
1961–1970 (1997).
15. Schlingmann, K. P. et al. Mutations in CYP24A1 and
idiopathic infantile hypercalcemia. N. Engl. J. Med.
365, 410–421 (2011).
16. Bikle, D. D., Siiteri, P. K., Ryzen, E. & Haddad, J. G.
Serum protein binding of 1,25-dihydroxyvitamin D:
a reevaluation by direct measurement of free
metabolite levels. J. Clin. Endocrinol. Metab. 61,
969–975 (1985).
17. Bikle, D. D. et al. Assessment of the free fraction
of 25-hydroxyvitamin D in serum and its regulation
by albumin and the vitamin D-binding protein.
J. Clin. Endocrinol. Metab. 63, 954–959 (1986).
18. Malik, S. et al. Common variants of the vitamin D
binding protein gene and adverse health outcomes.
Crit. Rev. Clin. Lab. Sci. 50, 1–22 (2013).
19. Bikle, D. D. & Schwartz, J. Vitamin D binding protein,
total and free vitamin D levels in different physiological
and pathophysiological conditions. Front. Endocrinol.
10, 317 (2019).
20. Bikle, D. D., Malmstroem, S. & Schwartz, J. Current
Controversies: are free vitamin metabolite levels a
more accurate assessment of vitamin D status than
total levels? Endocrinol. Metab. Clin. North Am. 46,
901–918 (2017).
21. Safadi, F. F. et al. Osteopathy and resistance to
vitamin D toxicity in mice null for vitamin D binding
protein. J. Clin. Invest. 103, 239–251 (1999).
Nature Reviews | Endocrinology
remains just that — speculation. We conclude our dis-
cussion by demonstrating two important outcomes of
vitamin D signalling in the skin: cancer suppression and
wound healing.
Published online xx xx xxxx
22. Henderson, C. M. et al. Vitamin D-binding protein
deficiency and homozygous deletion of the GC gene.
N. Engl. J. Med. 380, 1150–1157 (2019).
23. Wu, S. et al. Intracellular vitamin D binding proteins:
novel facilitators of vitamin D-directed transactivation.
Mol. Endocrinol. 14, 1387–1397 (2000).
24. Khanal, R. & Nemere, I. Membrane receptors for
vitamin D metabolites. Crit. Rev. Eukaryot. Gene Expr.
17, 31–47 (2007).
25. Pike, J. W., Meyer, M. B. & Bishop, K. A. Regulation of
target gene expression by the vitamin D receptor - an
update on mechanisms. Rev. Endocr. Metab. Disord.
13, 45–55 (2012).
26. Demay, M. B. The hair cycle and vitamin D receptor.
Arch. Biochem. Biophys. 523, 19–21 (2012).
27. Meyer, M. B. et al. A kidney-specific genetic control
module in mice governs endocrine regulation of
the cytochrome P450 gene Cyp27b1 essential for
vitamin D3 activation. J. Biol. Chem. 292,
17541–17558 (2017).
28. Meyer, M. B. et al. A chromatin-based mechanism
controls differential regulation of the cytochrome
P450 gene Cyp24a1 in renal and non-renal tissues.
J. Biol. Chem. 294, 14467–14481 (2019).
29. Meyer, M. B., Benkusky, N. A., Lee, C. H. & Pike, J. W.
Genomic determinants of gene regulation by
1,25-dihydroxyvitamin D3 during osteoblast-
lineage cell differentiation. J. Biol. Chem. 289,
19539–19554 (2014).
30. Malloy, P. J. et al. Vitamin D receptor mutations in
patients with hereditary 1,25-dihydroxyvitamin
D-resistant rickets. Mol. Genet. Metab. 111, 33–40
(2014).
31. Bikle, D. D. Vitamin D metabolism and function in the
skin. Mol. Cell. Endocrinol. 347, 80–89 (2011).
32. Barrea, L. et al. Vitamin D and its role in psoriasis:
an overview of the dermatologist and nutritionist.
Rev. Endocr. Metab. Disord. 18, 195–205 (2017).
33. Mitsche, M. A., McDonald, J. G., Hobbs, H. H. &
Cohen, J. C. Flux analysis of cholesterol biosynthesis
in vivo reveals multiple tissue and cell-type specific
pathways. eLife 4, e07999 (2015).
34. Prabhu, A. V., Luu, W., Sharpe, L. J. & Brown, A. J.
Cholesterol-mediated degradation of
7-dehydrocholesterol reductase switches the balance
from cholesterol to vitamin D synthesis. J. Biol. Chem.
291, 8363–8373 (2016).
35. Prabhu, A. V., Luu, W., Sharpe, L. J. & Brown, A. J.
Phosphorylation regulates activity of
7-dehydrocholesterol reductase (DHCR7), a terminal
enzyme of cholesterol synthesis. J. Steroid Biochem.
Mol. Biol. 165, 363–368 (2017).
36. Tint, G. S. et al. Defective cholesterol biosynthesis
associated with the Smith-Lemli-Opitz syndrome.
N. Engl. J. Med. 330, 107–113 (1994).
37. Xu, G. et al. Reproducing abnormal cholesterol
biosynthesis as seen in the Smith-Lemli-Opitz
syndrome by inhibiting the conversion of
7-dehydrocholesterol to cholesterol in rats.
J. Clin. Invest. 95, 76–81 (1995).
38. Movassaghi, M., Bianconi, S., Feinn, R., Wassif, C. A.
& Porter, F. D. Vitamin D levels in Smith-Lemli-Opitz
syndrome. Am. J. Med. Genet. A 173, 2577–2583
(2017).
39. Ahn, J. et al. Genome-wide association study of
circulating vitamin D levels. Hum. Mol. Genet. 19,
2739–2745 (2010).
40. Wang, T. J. et al. Common genetic determinants of
vitamin D insufficiency: a genome-wide association
study. Lancet 376, 180–188 (2010).
41. Kuan, V., Martineau, A. R., Griffiths, C. J., Hypponen, E.
& Walton, R. DHCR7 mutations linked to higher
vitamin D status allowed early human migration to
northern latitudes. BMC Evol. Biol. 13, 144 (2013).
42. Zhu, J. & DeLuca, H. F. Vitamin D 25-hydroxylase -
four decades of searching, are we there yet?
Arch. Biochem. Biophys. 523, 30–36 (2012).
43. Moghadasian, M. H. Cerebrotendinous
xanthomatosis: clinical course, genotypes and
metabolic backgrounds. Clin. Invest. Med. 27, 42–50
(2004).
Reviews
44. Cheng, J. B., Motola, D. L., Mangelsdorf, D. J. &
Russell, D. W. De-orphanization of cytochrome
P450 2R1: a microsomal vitamin D 25-hydroxylase.
J. Biol. Chem. 278, 38084–38093 (2003).
45. Al Mutair, A. N., Nasrat, G. H. & Russell, D. W.
Mutation of the CYP2R1 vitamin D 25-hydroxylase in a
Saudi Arabian family with severe vitamin D deficiency.
J. Clin. Endocrinol. Metab. 97, E2022–E2025 (2012).
46. Molin, A. et al. Vitamin D-dependent rickets type 1B
(25-hydroxylase deficiency): a rare condition or a
misdiagnosed condition? J. Bone Miner. Res. 32,
1893–1899 (2017).
47. Gupta, R. P., Hollis, B. W., Patel, S. B., Patrick, K. S. &
Bell, N. H. CYP3A4 is a human microsomal vitamin D
25-hydroxylase. J. Bone Miner. Res. 19, 680–688
(2004).
48. Gupta, R. P., He, Y. A., Patrick, K. S., Halpert, J. R.
& Bell, N. H. CYP3A4 is a vitamin D-24- and
25-hydroxylase: analysis of structure function by
site-directed mutagenesis. J. Clin. Endocrinol. Metab.
90, 1210–1219 (2005).
49. Roizen, J. D. et al. CYP3A4 mutation causes vitamin
D-dependent rickets type 3. J. Clin. Invest. 128,
1913–1918 (2018).
50. Adams, J. S. et al. Regulation of the extrarenal
CYP27B1-hydroxylase. J. Steroid Biochem. Mol. Biol.
144, 22–27 (2014).
51. Tebben, P. J., Singh, R. J. & Kumar, R. Vitamin
D-mediated hypercalcemia: mechanisms, diagnosis,
and treatment. Endocr. Rev. 37, 521–547 (2016).
52. Fraser, D. et al. Pathogenesis of hereditary vitamin-D-
dependent rickets — an inborn error of vitamin D
metabolism involving defective conversion of
25-hydroxyvitamin D to 1α,25-dihydroxyvitamin D.
N. Engl. J. Med. 289, 817–822 (1973).
53. Scriver, C. R., Reade, T. M., DeLuca, H. F. &
Hamstra, A. J. Serum 1,25-dihydroxyvitamin D levels
in normal subjects and in patients with hereditary
rickets or bone disease. N. Engl. J. Med. 299,
976–979 (1978).
54. Delvin, E. E., Glorieux, F. H., Marie, P. J. & Pettifor, J. M.
Vitamin D dependency: replacement therapy with
calcitriol? J. Pediatr. 99, 26–34 (1981).
55. St-Arnaud, R., Messerlian, S., Moir, J. M., Omdahl, J. L.
& Glorieux, F. H. The 25-hydroxyvitamin D
1-alpha-hydroxylase gene maps to the pseudovitamin
D-deficiency rickets (PDDR) disease locus. J. Bone
Miner. Res. 12, 1552–1559 (1997).
56. Takeyama, K. et al. 25-Hydroxyvitamin D3
1alpha-hydroxylase and vitamin D synthesis.
Science 277, 1827–1830 (1997).
57. Monkawa, T. et al. Molecular cloning of cDNA and
genomic DNA for human 25-hydroxyvitamin
D31α-hydroxylase. Biochem. Biophys. Res. Commun.
239, 527–533 (1997).
58. Kim, C. J. et al. Vitamin D 1α-hydroxylase gene
mutations in patients with 1α-hydroxylase deficiency.
J. Clin. Endocrinol. Metab. 92, 3177–3182 (2007).
59. Glorieux, F. H. & Pettifor, J. M. Vitamin D/dietary
calcium deficiency rickets and pseudo-vitamin D
deficiency rickets. Bonekey Rep. 3, 524 (2014).
60. Bitzan, M. & Goodyer, P. R. Hypophosphatemic
rickets. Pediatr. Clin. North Am. 66, 179–207
(2019).
61. Xu, Y. et al. Intestinal and hepatic CYP3A4 catalyze
hydroxylation of 1α,25-dihydroxyvitamin D3:
implications for drug-induced osteomalacia.
Mol. Pharmacol. 69, 56–65 (2006).
62. Prosser, D. E., Kaufmann, M., O'Leary, B., Byford, V.
& Jones, G. Single A326G mutation converts human
CYP24A1 from 25-OH-D3-24-hydroxylase into
-23-hydroxylase, generating 1α,25-(OH)2D3-26,
23-lactone. Proc. Natl Acad. Sci. USA 104,
12673–12678 (2007).
63. Jones, G., Prosser, D. E. & Kaufmann, M.
25-Hydroxyvitamin D-24-hydroxylase (CYP24A1):
its important role in the degradation of vitamin D.
Arch. Biochem. Biophys. 523, 9–18 (2012).
64. Brodie, M. J. et al. Effect of rifampicin and isoniazid
on vitamin D metabolism. Clin. Pharmacol. Ther. 32,
525–530 (1982).
Reviews
65. Sakaki, T. et al. Dual metabolic pathway of
25-hydroxyvitamin D3 catalyzed by human CYP24.
Eur. J. Biochem. 267, 6158–6165 (2000).
66. Harant, H., Spinner, D., Reddy, G. S. & Lindley, I. J.
Natural metabolites of 1α,25-dihydroxyvitamin D3
retain biologic activity mediated through the vitamin D
receptor. J. Cell. Biochem. 78, 112–120 (2000).
67. Martineau, C. et al. Optimal bone fracture repair
requires 24R,25-dihydroxyvitamin D3 and its effector
molecule FAM57B2. J. Clin. Invest. 128, 3546–3557
(2018).
68. Plachot, J. J. et al. In vitro action of
1,25-dihydroxycholecalciferol and 24,25-
dihydroxycholecalciferol on matrix organization and
mineral distribution in rabbit growth plate.
Metab. Bone Dis. Relat. Res. 4, 135–142 (1982).
69. St-Arnaud, R. et al. Deficient mineralization of
intramembranous bone in vitamin D-24-hydroxylase-
ablated mice is due to elevated 1,25-dihydroxyvitamin D
and not to the absence of 24,25-dihydroxyvitamin D.
Endocrinology 141, 2658–2666 (2000).
70. Jones, G., Kottler, M. L. & Schlingmann, K. P. Genetic
diseases of vitamin D metabolizing enzymes.
Endocrinol. Metab. Clin. North Am. 46, 1095–1117
(2017).
71. Dinour, D. et al. Maternal and infantile hypercalcemia
caused by vitamin-D-hydroxylase mutations and
vitamin D intake. Pediatr. Nephrol. 30, 145–152
(2015).
72. Shah, A. D. et al. Maternal hypercalcemia due to
failure of 1,25-dihydroxyvitamin-D3 catabolism in a
patient with CYP24A1 mutations. J. Clin. Endocrinol.
Metab. 100, 2832–2836 (2015).
73. Tebben, P. J. et al. Hypercalcemia, hypercalciuria, and
elevated calcitriol concentrations with autosomal
dominant transmission due to CYP24A1 mutations:
effects of ketoconazole therapy. J. Clin. Endocrinol.
Metab. 97, E423–E427 (2012).
74. Cools, M. et al. Calcium and bone homeostasis in
heterozygous carriers of CYP24A1 mutations:
a cross-sectional study. Bone 81, 89–96 (2015).
75. Molin, A. et al. CYP24A1 mutations in a cohort of
hypercalcemic patients: evidence for a recessive trait.
J. Clin. Endocrinol. Metab. 100, E1343–E1352 (2015).
76. Lightwood, R. & Stapleton, T. Idiopathic hypercalcaemia
in infants. Lancet 265, 255–256 (1953).
77. Pober, B. R. Williams-Beuren syndrome. N. Engl.
J. Med. 362, 239–252 (2010).
78. Chen, A. et al. Description of 5 novel SLC34A3/NPT2c
mutations causing hereditary hypophosphatemic
rickets with hypercalciuria. Kidney Int. Rep. 4,
1179–1186 (2019).
79. Bergwitz, C. & Miyamoto, K. I. Hereditary
hypophosphatemic rickets with hypercalciuria:
pathophysiology, clinical presentation, diagnosis and
therapy. Pflugers Arch. 471, 149–163 (2019).
80. Kaufmann, M. et al. Clinical utility of simultaneous
quantitation of 25-hydroxyvitamin D and
24,25-dihydroxyvitamin D by LC-MS/MS involving
derivatization with DMEQ-TAD. J. Clin. Endocrinol.
Metab. 99, 2567–2574 (2014).
81. Hirschfeld, J. Immune-electrophoretic demonstration
of qualitative differences in human sera and their
relation to the haptoglobins. Acta Pathol. Microbiol.
Scand. 47, 160–168 (1959).
82. Daiger, S. P., Schanfield, M. S. & Cavalli-Sforza, L. L.
Group-specific component (Gc) proteins bind vitamin D
and 25-hydroxyvitamin D. Proc. Natl Acad. Sci. USA
72, 2076–2080 (1975).
83. Armas, L. A., Hollis, B. W. & Heaney, R. P. Vitamin D2
is much less effective than vitamin D3 in humans.
J. Clin. Endocrinol. Metab. 89, 5387–5391 (2004).
84. Mendel, C. M. The free hormone hypothesis:
a physiologically based mathematical model.
Endocr. Rev. 10, 232–274 (1989).
85. Nykjaer, A. et al. An endocytic pathway essential for
renal uptake and activation of the steroid 25-(OH)
vitamin D3. Cell 96, 507–515 (1999).
86. Zella, L. A., Shevde, N. K., Hollis, B. W., Cooke, N. E.
& Pike, J. W. Vitamin D-binding protein influences
total circulating levels of 1,25-dihydroxyvitamin D3
but does not directly modulate the bioactive levels of
the hormone in vivo. Endocrinology 149, 3656–3667
(2008).
87. Cooke, N. E., McLeod, J. F., Wang, X. K. & Ray, K.
Vitamin D binding protein: genomic structure,
functional domains, and mRNA expression in tissues.
J. Steroid Biochem. Mol. Biol. 40, 787–793 (1991).
88. Hagenfeldt, Y., Carlstrom, K., Berlin, T. & Stege, R.
Effects of orchidectomy and different modes of high
dose estrogen treatment on circulating “free” and total
1,25-dihydroxyvitamin D in patients with prostatic
cancer. J. Steroid Biochem. Mol. Biol. 39, 155–159
(1991).
89. Moller, U. K., Streym, S., Heickendorff, L., Mosekilde, L.
& Rejnmark, L. Effects of 25OHD concentrations on
chances of pregnancy and pregnancy outcomes: a
cohort study in healthy Danish women. Eur. J. Clin.
Nutr. 66, 862–868 (2012).
90. Zhang, J. Y., Lucey, A. J., Horgan, R., Kenny, L. C. &
Kiely, M. Impact of pregnancy on vitamin D status:
a longitudinal study. Br. J. Nutr. 112, 1081–1087
(2014).
91. Moller, U. K. et al. Increased plasma concentrations
of vitamin D metabolites and vitamin D binding
protein in women using hormonal contraceptives:
a cross-sectional study. Nutrients 5, 3470–3480
(2013).
92. Guha, C., Osawa, M., Werner, P. A., Galbraith, R. M. &
Paddock, G. V. Regulation of human Gc (vitamin D–
binding) protein levels: hormonal and cytokine control of
gene expression in vitro. Hepatology 21, 1675–1681
(1995).
93. Dahl, B. et al. Trauma stimulates the synthesis of
Gc-globulin. Intensive Care Med. 27, 394–399
(2001).
94. Schiodt, F. V. Gc-globulin in liver disease. Dan. Med.
Bull. 55, 131–146 (2008).
95. Wang, X., Shapses, S. A. & Al-Hraishawi, H. Free and
bioavailable 25-hydroxyvitamin D levels in patients
with primary hyperparathyroidism. Endocr. Pract. 23,
66–71 (2017).
96. Song, Y. H., Ray, K., Liebhaber, S. A. & Cooke, N. E.
Vitamin D-binding protein gene transcription is
regulated by the relative abundance of hepatocyte
nuclear factors 1α and 1β. J. Biol. Chem. 273,
28408–28418 (1998).
97. Cleve, H. & Constans, J. The mutants of the vitamin-D-
binding protein: more than 120 variants of the GC/
DBP system. Vox Sang. 54, 215–225 (1988).
98. Chun, R. F. New perspectives on the vitamin D binding
protein. Cell Biochem. Funct. 30, 445–456 (2012).
99. Nagasawa, H. et al. Gc protein (vitamin D-binding
protein): Gc genotyping and GcMAF precursor activity.
Anticancer Res. 25, 3689–3695 (2005).
100. Uto, Y. et al. β-Galactosidase treatment is a common
first-stage modification of the three major subtypes
of Gc protein to GcMAF. Anticancer Res. 32,
2359–2364 (2012).
101. Arnaud, J. & Constans, J. Affinity differences for
vitamin D metabolites associated with the genetic
isoforms of the human serum carrier protein (DBP).
Hum. Genet. 92, 183–188 (1993).
102. Bouillon, R., van Baelen, H. & de Moor, P. Comparative
study of the affinity of the serum vitamin D-binding
protein. J. Steroid Biochem. 13, 1029–1034 (1980).
103. Boutin, B., Galbraith, R. M. & Arnaud, P. Comparative
affinity of the major genetic variants of human
group-specific component (vitamin D-binding protein)
for 25-(OH) vitamin D. J. Steroid Biochem. 32, 59–63
(1989).
104. Jones, K. S. et al. 25(OH)D2 half-life is shorter than
25(OH)D3 half-life and is influenced by DBP
concentration and genotype. J. Clin. Endocrinol. Metab.
99, 3373–3381 (2014).
105. Chun, R. F. et al. Vitamin D-binding protein directs
monocyte responses to 25-hydroxy- and
1,25-dihydroxyvitamin D. J. Clin. Endocrinol. Metab.
95, 3368–3376 (2010).
106. Bikle, D. D., Gee, E., Halloran, B. & Haddad, J. G.
Free 1,25-dihydroxyvitamin D levels in serum from
normal subjects, pregnant subjects, and subjects
with liver disease. J. Clin. Invest. 74, 1966–1971
(1984).
107. Schwartz, J. B. et al. A comparison of measured and
calculated free 25(OH) vitamin D levels in clinical
populations. J. Clin. Endocrinol. Metab. 99,
1631–1637 (2014).
108. Bikle, D. D., Halloran, B. P., Gee, E., Ryzen, E. &
Haddad, J. G. Free 25-hydroxyvitamin D levels are
normal in subjects with liver disease and reduced
total 25-hydroxyvitamin D levels. J. Clin. Invest. 78,
748–752 (1986).
109. Schwartz, J. B. et al. Determination of free 25(OH)D
concentrations and their relationships to total 25(OH)
D in multiple clinical populations. J. Clin. Endocrinol.
Metab. 103, 3278–3288 (2018).
110. Schwartz, J. B., Kane, L. & Bikle, D. Response of
vitamin D concentration to vitamin D3 administration
in older adults without sun exposure: a randomized
double-blind trial. J. Am. Geriatr. Soc. 64, 65–72
(2016).
111. Lauridsen, A. L., Vestergaard, P. & Nexo, E. Mean
serum concentration of vitamin D-binding protein
(Gc globulin) is related to the Gc phenotype in women.
Clin. Chem. 47, 753–756 (2001).
112. Hoofnagle, A. N., Eckfeldt, J. H. & Lutsey, P. L.
Vitamin D-binding protein concentrations quantified
by mass spectrometry. N. Engl. J. Med. 373,
1480–1482 (2015).
113. Carpenter, T. O. et al. Vitamin D binding protein is a
key determinant of 25-hydroxyvitamin D levels in
infants and toddlers. J. Bone Miner. 28, 213–221
(2013).
114. Santos, B. R., Mascarenhas, L. P., Boguszewski, M. C. &
Spritzer, P. M. Variations in the vitamin D-binding protein
(DBP) gene are related to lower 25-hydroxyvitamin D
levels in healthy girls: a cross-sectional study. Horm. Res.
Paediatr. 79, 162–168 (2013).
115. Sollid, S. T. et al. Effects of vitamin D binding protein
phenotypes and vitamin D supplementation on serum
total 25(OH)D and directly measured free 25(OH)D.
Eur. J. Endocrinol 174, 445–452 (2016).
116. Takiar, R. et al. The associations of 25-hydroxyvitamin D
levels, vitamin D binding protein gene polymorphisms,
and race with risk of incident fracture-related
hospitalization: twenty-year follow-up in a bi-ethnic
cohort (the ARIC study). Bone 78, 94–101 (2015).
117. Leong, A. et al. The causal effect of vitamin D binding
protein (DBP) levels on calcemic and cardiometabolic
diseases: a Mendelian randomization study.
PLOS Med. 11, e1001751 (2014).
118. Hirai, M. et al. Group specific component protein
genotype is associated with NIDDM in Japan.
Diabetologia 41, 742–743 (1998).
119. Baier, L. J., Dobberfuhl, A. M., Pratley, R. E.,
Hanson, R. L. & Bogardus, C. Variations in the vitamin
D-binding protein (Gc locus) are associated with oral
glucose tolerance in nondiabetic Pima Indians. J. Clin.
Endocrinol. Metab. 83, 2993–2996 (1998).
120. Ye, W. Z., Dubois-Laforgue, D., Bellanne-Chantelot, C.,
Timsit, J. & Velho, G. Variations in the vitamin
D-binding protein (Gc locus) and risk of type 2
diabetes mellitus in French Caucasians. Metabolism
50, 366–369 (2001).
121. Lauridsen, A. L. et al. Female premenopausal fracture
risk is associated with Gc phenotype. J. Bone Miner.
19, 875–881 (2004).
122. Papiha, S. S., Allcroft, L. C., Kanan, R. M., Francis, R. M.
& Datta, H. K. Vitamin D binding protein gene in male
osteoporosis: association of plasma DBP and bone
mineral density with (TAAA)(n)-Alu polymorphism in
DBP. Calcif. Tissue Int. 65, 262–266 (1999).
123. Ezura, Y. et al. Association of molecular variants,
haplotypes, and linkage disequilibrium within the
human vitamin D-binding protein (DBP) gene with
postmenopausal bone mineral density. J. Bone Miner.
Res. 18, 1642–1649 (2003).
124. Chishimba, L., Thickett, D. R., Stockley, R. A. &
Wood, A. M. The vitamin D axis in the lung: a key role
for vitamin D-binding protein. Thorax 65, 456–462
(2010).
125. Faserl, K. et al. Polymorphism in vitamin D-binding
protein as a genetic risk factor in the pathogenesis
of endometriosis. J. Clin. Endocrinol. Metab. 96,
E233–E241 (2011).
126. Eloranta, J. J. et al. Association of a common vitamin
D-binding protein polymorphism with inflammatory
bowel disease. Pharmacogenet Genomics 21, 559–564
(2011).
127. Abbas, S. et al. The Gc2 allele of the vitamin D binding
protein is associated with a decreased postmenopausal
breast cancer risk, independent of the vitamin D status.
Cancer Epidemiol. Biomarkers Prev. 17, 1339–1343
(2008).
128. Dimopoulos, M. A. et al. Genetic markers in carcinoma
of the prostate. Eur. Urol. 10, 315–316 (1984).
129. Zhou, L. et al. GC Glu416Asp and Thr420Lys
polymorphisms contribute to gastrointestinal cancer
susceptibility in a Chinese population. Int. J. Clin.
Exp. Med. 5, 72–79 (2012).
130. Poynter, J. N. et al. Genetic variation in the vitamin D
receptor (VDR) and the vitamin D-binding protein (GC)
and risk for colorectal cancer: results from the Colon
Cancer Family Registry. Cancer Epidemiol. Biomarkers
Prev. 19, 525–536 (2010).
131. Martineau, A. R. et al. Association between Gc
genotype and susceptibility to TB is dependent on
vitamin D status. Eur. Respir. J. 35, 1106–1112 (2010).
132. Gacad, M. A., Chen, H., Arbelle, J. E., LeBon, T.
& Adams, J. S. Functional characterization and
purification of an intracellular vitamin D-binding
protein in vitamin D-resistant New World primate
cells. Amino acid sequence homology with proteins
in the hsp-70 family. J. Biol. Chem. 272, 8433–8440
(1997).
www.nature.com/nrendo

133. Adams, J. S. et al. Response element binding proteins
and intracellular vitamin D binding proteins: novel
regulators of vitamin D trafficking, action and
metabolism. J. Steroid Biochem. Mol. Biol. 89-90,
461–465 (2004).
134. Brumbaugh, P. F. & Haussler, M. R. 1α,25-
Dihydroxycholecalciferol receptors in intestine.
II. Temperature-dependent transfer of the hormone to
chromatin via a specific cytosol receptor. J. Biol. Chem.
249, 1258–1262 (1974).
135. Wang, Y., Zhu, J. & DeLuca, H. F. Where is the vitamin D
receptor? Arch. Biochem. Biophys. 523, 123–133
(2012).
136. Christakos, S., Dhawan, P., Verstuyf, A., Verlinden, L.
& Carmeliet, G. Vitamin D: metabolism, molecular
mechanism of action, and pleiotropic effects.
Physiol. Rev. 96, 365–408 (2016).
137. Gunton, J. E. & Girgis, C. M. Vitamin D and muscle.
Bone Rep. 8, 163–167 (2018).
138. Zheng, J. et al. HDX reveals the conformational
dynamics of DNA sequence specific VDR co-activator
interactions. Nat. Commun. 8, 923 (2017).
139. Shaffer, P. L. & Gewirth, D. T. Structural basis of
VDR-DNA interactions on direct repeat response
elements. EMBO J. 21, 2242–2252 (2002).
140. Rochel, N., Wurtz, J. M., Mitschler, A., Klaholz, B. &
Moras, D. The crystal structure of the nuclear receptor
for vitamin D bound to its natural ligand. Mol. Cell 5,
173–179 (2000).
141. Carlberg, C., Molnár, F. & Mouriño, A. Vitamin D
receptor ligands: the impact of crystal structures.
Expert. Opin. Ther. Pat. 22, 417–435 (2012).
142. Mizwicki, M. T. & Norman, A. W. The vitamin D
sterol-vitamin D receptor ensemble model offers
unique insights into both genomic and rapid-response
signaling. Sci. Signal. 2, re4 (2009).
143. Singarapu, K. K. et al. Ligand-specific structural
changes in the vitamin D receptor in solution.
Biochemistry 50, 11025–11033 (2011).
144. Rochel, N. et al. Common architecture of nuclear
receptor heterodimers on DNA direct repeat elements
with different spacings. Nat. Struct. Mol. Biol. 18,
564–570 (2011).
145. Orlov, I., Rochel, N., Moras, D. & Klaholz, B. P.
Structure of the full human RXR/VDR nuclear receptor
heterodimer complex with its DR3 target DNA.
EMBO J. 31, 291–300 (2012).
146. Zhang, J. et al. DNA binding alters coactivator
interaction surfaces of the intact VDR-RXR complex.
Nat. Struct. Mol. Biol. 18, 556–563 (2011).
147. Feldman, D. & Malloy, P. J. Mutations in the vitamin D
receptor and hereditary vitamin D-resistant rickets.
Bonekey Rep. 3, 510 (2014).
148. Lee, S. M., Goellner, J. J., O'Brien, C. A. & Pike, J. W.
A humanized mouse model of hereditary
1,25-dihydroxyvitamin D-resistant rickets without
alopecia. Endocrinology 155, 4137–4148 (2014).
149. Huet, T. et al. A vitamin D receptor selectively activated
by gemini analogs reveals ligand dependent and
independent effects. Cell Rep. 10, 516–526 (2015).
150. Lee, S. M. & Pike, J. W. The vitamin D receptor
functions as a transcription regulator in the absence of
1,25-dihydroxyvitamin D3. J. Steroid Biochem. Mol.
Biol. 164, 265–270 (2016).
151. Balsan, S. et al. Long-term nocturnal calcium infusions
can cure rickets and promote normal mineralization
in hereditary resistance to 1,25-dihydroxyvitamin D.
J. Clin. Invest. 77, 1661–1667 (1986).
152. Chaturvedi, D., Garabedian, M., Carel, J. C. & Leger, J.
Different mechanisms of intestinal calcium absorption
at different life stages: therapeutic implications and
long-term responses to treatment in patients with
hereditary vitamin D-resistant rickets. Horm. Res.
Paediatr. 78, 326–331 (2012).
153. Pike, J. W. & Christakos, S. Biology and mechanisms of
action of the vitamin D hormone. Endocrinol. Metab.
Clin. North Am. 46, 815–843 (2017).
154. Pike, J. W., Meyer, M. B., Lee, S. M., Onal, M. &
Benkusky, N. A. The vitamin D receptor: contemporary
genomic approaches reveal new basic and
translational insights. J. Clin. Invest. 127, 1146–1154
(2017).
155. Wei, R., Dhawan, P., Baiocchi, R. A., Kim, K. Y. &
Christakos, S. PU.1 and epigenetic signals modulate
1,25-dihydroxyvitamin D3 and C/EBPα regulation
of the human cathelicidin antimicrobial peptide
gene in lung epithelial cells. J. Cell Physiol. 234,
10345–10359 (2019).
156. Seth-Vollenweider, T., Joshi, S., Dhawan, P., Sif, S.
& Christakos, S. Novel mechanism of negative
regulation of 1,25-dihydroxyvitamin D3-induced
25-hydroxyvitamin D3 24-hydroxylase (Cyp24a1)
Nature Reviews | Endocrinology
Transcription: epigenetic modification involving
cross-talk between protein-arginine methyltransferase
5 and the SWI/SNF complex. J. Biol. Chem. 289,
33958–33970 (2014).
157. Meyer, M. B., Benkusky, N. A. & Pike, J. W.
The RUNX2 cistrome in osteoblasts: characterization,
down-regulation following differentiation, and
relationship to gene expression. J. Biol. Chem. 289,
16016–16031 (2014).
158. Pike, J. W., Meyer, M. B., St John, H. C. &
Benkusky, N. A. Epigenetic histone modifications
and master regulators as determinants of context
dependent nuclear receptor activity in bone cells.
Bone 81, 757–764 (2015).
159. Ding, N. et al. A vitamin D receptor/SMAD genomic
circuit gates hepatic fibrotic response. Cell 153,
601–613 (2013).
160. Sherman, M. H. et al. Vitamin D receptor-mediated
stromal reprogramming suppresses pancreatitis and
enhances pancreatic cancer therapy. Cell 159, 80–93
(2014).
161. Wei, Z. et al. Vitamin D switches BAF complexes to
protect β cells. Cell 173, 1135–1149.e15 (2018).
162. Bikle, D. D., Nemanic, M. K., Whitney, J. O. & Elias, P. W.
Neonatal human foreskin keratinocytes produce
1,25-dihydroxyvitamin D3. Biochemistry 25,
1545–1548 (1986).
163. Stumpf, W. E., Sar, M., Reid, F. A., Tanaka, Y. &
DeLuca, H. F. Target cells for 1,25-dihydroxyvitamin D3
in intestinal tract, stomach, kidney, skin, pituitary, and
parathyroid. Science 206, 1188–1190 (1979).
164. Pillai, S., Bikle, D. D. & Elias, P. M. 1,25-
Dihydroxyvitamin D production and receptor binding
in human keratinocytes varies with differentiation.
J. Biol. Chem. 263, 5390–5395 (1988).
165. Hosomi, J., Hosoi, J., Abe, E., Suda, T. & Kuroki, T.
Regulation of terminal differentiation of cultured
mouse epidermal cells by 1α,25-dihydroxyvitamin D3.
Endocrinology 113, 1950–1957 (1983).
166. Smith, E. L., Walworth, N. C. & Holick, M. F. Effect of
1α,25-dihydroxyvitamin D3 on the morphologic and
biochemical differentiation of cultured human
epidermal keratinocytes grown in serum-free
conditions. J. Invest. Dermatol. 86, 709–714 (1986).
167. Oda, Y. et al. Vitamin D receptor and coactivators
SRC2 and 3 regulate epidermis-specific sphingolipid
production and permeability barrier formation.
J. Invest. Dermatol. 129, 1367–1378 (2009).
168. Tu, C. L. et al. Ablation of the calcium-sensing receptor
in keratinocytes impairs epidermal differentiation and
barrier function. J. Invest. Dermatol. 132, 2350–2359
(2012).
169. Oda, Y. et al. Vitamin D receptor is required for
proliferation, migration, and differentiation of
epidermal stem cells and progeny during cutaneous
wound repair. J. Invest. Dermatol. 138, 2423–2431
(2018).
170. Tu, C. L., Celli, A., Mauro, T. & Chang, W. The
calcium-sensing receptor regulates epidermal
intracellular Ca(2+) signaling and re-epithelialization
after wounding. J. Invest. Dermatol. 139, 919–929
(2019).
171. Teichert, A., Elalieh, H. & Bikle, D. Disruption of the
hedgehog signaling pathway contributes to the
hair follicle cycling deficiency in Vdr knockout mice.
J. Cell. Physiol. 225, 482–489 (2010).
172. Bikle, D. D., Elalieh, H., Chang, S., Xie, Z. &
Sundberg, J. P. Development and progression of
alopecia in the vitamin D receptor null mouse. J. Cell.
Physiol. 207, 340–353 (2006).
173. Li, Y. C. et al. Targeted ablation of the vitamin D
receptor: an animal model of vitamin D-dependent
rickets type II with alopecia. Proc. Natl Acad. Sci. USA
94, 9831–9835 (1997).
174. Palmer, H. G., Anjos-Afonso, F., Carmeliet, G.,
Takeda, H. & Watt, F. M. The vitamin D receptor is a
Wnt effector that controls hair follicle differentiation
and specifies tumor type in adult epidermis.
PLOS ONE 3, e1483 (2008).
175. Kretzschmar, K. & Watt, F. M. Markers of epidermal
stem cell subpopulations in adult mammalian skin.
Cold Spring Harb. Perspect. Med. 4, a013631
(2014).
176. Eichner, R., Sun, T. T. & Aebi, U. The role of keratin
subfamilies and keratin pairs in the formation of
human epidermal intermediate filaments. J. Cell Biol.
102, 1767–1777 (1986).
177. Warhol, M. J., Roth, J., Lucocq, J. M., Pinkus, G. S.
& Rice, R. H. Immuno-ultrastructural localization of
involucrin in squamous epithelium and cultured
keratinocytes. J. Histochem. Cytochem. 33, 141–149
(1985).
Reviews
178. Thacher, S. M. & Rice, R. H. Keratinocyte-specific
transglutaminase of cultured human epidermal cells:
relation to cross-linked envelope formation and
terminal differentiation. Cell 40, 685–695 (1985).
179. Steven, A. C., Bisher, M. E., Roop, D. R. & Steinert, P. M.
Biosynthetic pathways of filaggrin and loricrin–two
major proteins expressed by terminally differentiated
epidermal keratinocytes. J. Struct. Biol. 104, 150–162
(1990).
180. Freeman, S. C. & Sonthalia, S. Histology, keratohyalin
granules. StatPearls https://www.ncbi.nlm.nih.gov/
books/NBK537049/ (2019).
181. Dale, B. A., Resing, K. A. & Lonsdale-Eccles, J. D.
Filaggrin: a keratin filament associated protein.
Ann. NY Acad. Sci. 455, 330–342 (1985).
182. Levin, J., Friedlander, S. F. & Del Rosso, J. Q. Atopic
dermatitis and the stratum corneum: part 2: other
structural and functional characteristics of the stratum
corneum barrier in atopic skin. J. Clin. Aesthet.
Dermatol. 6, 49–54 (2013).
183. Mehrel, T. et al. Identification of a major keratinocyte
cell envelope protein, loricrin. Cell 61, 1103–1112
(1990).
184. Menon, G. K., Lee, S. E. & Lee, S. H. An overview of
epidermal lamellar bodies: novel roles in biological
adaptations and secondary barriers. J. Dermatol. Sci.
92, 10–17 (2018).
185. Feingold, K. R. & Elias, P. M. Role of lipids in the
formation and maintenance of the cutaneous
permeability barrier. Biochim. Biophys. Acta 1841,
280–294 (2014).
186. Aberg, K. M. et al. Co-regulation and interdependence
of the mammalian epidermal permeability and
antimicrobial barriers. J. Invest. Dermatol. 128,
917–925 (2008).
187. Bikle, D. D. & Pillai, S. Vitamin D, calcium, and epidermal
differentiation. Endocr. Rev. 14, 3–19 (1993).
188. Bikle, D. D., Pillai, S. & Gee, E. Squamous carcinoma
cell lines produce 1,25 dihydroxyvitamin D, but fail to
respond to its prodifferentiating effect. J. Invest.
Dermatol. 97, 435–441 (1991).
189. McLane, J. A., Katz, M. & Abdelkader, N. Effect of
1,25-dihydroxyvitamin D3 on human keratinocytes
grown under different culture conditions. In Vitro Cell.
Dev. Biol. 26, 379–387 (1990).
190. Hawker, N. P., Pennypacker, S. D., Chang, S. M.
& Bikle, D. D. Regulation of human epidermal
keratinocyte differentiation by the vitamin D receptor
and its coactivators DRIP205, SRC2, and SRC3.
J. Invest. Dermatol. 127, 874–880 (2007).
191. Schauber, J. et al. Injury enhances TLR2 function and
antimicrobial peptide expression through a vitamin
D-dependent mechanism. J. Clin. Invest. 117, 803–811
(2007).
192. Schauber, J., Dorschner, R. A., Yamasaki, K., Brouha, B.
& Gallo, R. L. Control of the innate epithelial
antimicrobial response is cell-type specific and
dependent on relevant microenvironmental stimuli.
Immunology 118, 509–519 (2006).
193. Oda, Y. et al. Vitamin D receptor and coactivators
SRC 2 and 3 regulate epidermis-specific sphingolipid
production and permeability barrier formation.
J. Invest. Dermatol. 129, 1367–1378 (2009).
194. Muehleisen, B. et al. PTH/PTHrP and vitamin D
control antimicrobial peptide expression and
susceptibility to bacterial skin infection. Sci. Transl
Med. 4, 135ra66 (2012).
195. Zehnder, D. et al. Extrarenal expression of
25-hydroxyvitamin D3-1α-hydroxylase. J. Clin.
Endocrinol. Metab. 86, 888–894 (2001).
196. Stumpf, W. E., Clark, S. A., Sar, M. & DeLuca, H. F.
Topographical and developmental studies on target
sites of 1,25 (OH)2 vitamin D3 in skin. Cell Tissue Res.
238, 489–496 (1984).
197. Oda, Y. et al. Two distinct coactivators, DRIP/mediator
and SRC/p160, are differentially involved in vitamin D
receptor transactivation during keratinocyte
differentiation. Mol. Endocrinol. 17, 2329–2339
(2003).
198. Oda, Y., Ishikawa, M. H., Hawker, N. P., Yun, Q. C. &
Bikle, D. D. Differential role of two VDR coactivators,
DRIP205 and SRC-3, in keratinocyte proliferation and
differentiation. J. Steroid Biochem. Mol. Biol. 103,
776–780 (2007).
199. Oda, Y. et al. Coactivator MED1 ablation in
keratinocytes results in hair-cycling defects and
epidermal alterations. J. Invest. Dermatol. 132,
1075–1083 (2012).
200. Schauber, J. et al. Histone acetylation in keratinocytes
enables control of the expression of cathelicidin
and CD14 by 1,25-dihydroxyvitamin D3. J. Invest.
Dermatol. 128, 816–824 (2008).
Reviews
201. Bikle, D. D., Xie, Z. & Tu, C. L. Calcium regulation of
keratinocyte differentiation. Expert. Rev. Endocrinol.
Metab. 7, 461–472 (2012).
202. Tu, C. L., Chang, W. & Bikle, D. D. The extracellular
calcium-sensing receptor is required for
calcium-induced differentiation in human
keratinocytes. J. Biol. Chem. 276, 41079–41085
(2001).
203. Tu, C. L., Chang, W. & Bikle, D. D. Phospholipase Cγ1
is required for activation of store-operated channels
in human keratinocytes. J. Invest. Dermatol. 124,
187–197 (2005).
204. Tu, C., Chang, W., Xie, Z. & Bikle, D. Inactivation of the
calcium sensing receptor inhibits E-cadherin-mediated
cell-cell adhesion and calcium-induced differentiation
in human epidermal keratinocytes. J. Biol. Chem. 283,
3519–3528 (2008).
205. Xie, Z., Singleton, P. A., Bourguignon, L. Y. & Bikle, D. D.
Calcium-induced human keratinocyte differentiation
requires src- and fyn-mediated phosphatidylinositol
3-kinase-dependent activation of phospholipase C-γ1.
Mol. Biol. Cell 16, 3236–3246 (2005).
206. Xie, Z. & Bikle, D. D. The recruitment of
phosphatidylinositol 3-kinase to the E-cadherin-
catenin complex at the plasma membrane is required
for calcium-induced phospholipase C-γ1 activation and
human keratinocyte differentiation. J. Biol. Chem.
282, 8695–8703 (2007).
207. Xie, Z., Chang, S. M., Pennypacker, S. D., Liao, E. Y. &
Bikle, D. D. Phosphatidylinositol-4-phosphate 5-kinase
1α mediates extracellular calcium-induced keratinocyte
differentiation. Mol. Biol. Cell 20, 1695–1704 (2009).
208. Tu, C. L., Oda, Y., Komuves, L. & Bikle, D. D. The role
of the calcium-sensing receptor in epidermal
differentiation. Cell Calcium 35, 265–273 (2004).
209. Tu, C. L., Chang, W. & Bikle, D. D. The role of
the calcium sensing receptor in regulating intracellular
calcium handling in human epidermal keratinocytes.
J. Invest. Dermatol. 127, 1074–1083 (2007).
210. Canaff, L. & Hendy, G. N. Human calcium-sensing
receptor gene. Vitamin D response elements in
promoters P1 and P2 confer transcriptional
responsiveness to 1,25-dihydroxyvitamin D. J. Biol.
Chem. 277, 30337–30350 (2002).
211. Xie, Z. & Bikle, D. D. Cloning of the human
phospholipase C-γ1 promoter and identification of a
DR6-type vitamin D-responsive element. J. Biol. Chem.
272, 6573–6577 (1997).
212. Su, M. J., Bikle, D. D., Mancianti, M. L. & Pillai, S.
1,25-Dihydroxyvitamin D3 potentiates the keratinocyte
response to calcium. J. Biol. Chem. 269, 14723–14729
(1994).
213. Gniadecki, R., Gajkowska, B. & Hansen, M.
1,25-Dihydroxyvitamin D3 stimulates the assembly of
adherens junctions in keratinocytes: involvement of
protein kinase C. Endocrinology 138, 2241–2248
(1997).
214. Hu, L., Bikle, D. D. & Oda, Y. Reciprocal role of
vitamin D receptor on β-catenin regulated keratinocyte
proliferation and differentiation. J. Steroid Biochem.
Mol. Biol. 144, 237–241 (2014).
215. Xie, Z. et al. Lack of the vitamin D receptor is
associated with reduced epidermal differentiation and
hair follicle growth. J. Invest. Dermatol. 118, 11–16
(2002).
216. Watt, F. M. & Collins, C. A. Role of β-catenin in
epidermal stem cell expansion, lineage selection,
and cancer. Cold Spring Harb. Symp. Quant. Biol. 73,
503–512 (2008).
217. Luderer, H. F., Gori, F. & Demay, M. B. Lymphoid
enhancer-binding factor-1 (LEF1) interacts with
the DNA-binding domain of the vitamin D receptor.
J. Biol. Chem. 286, 18444–18451 (2011).
218. Hill, N. T. et al. 1α, 25-Dihydroxyvitamin D(3) and the
vitamin D receptor regulates ΔNp63α levels and
keratinocyte proliferation. Cell Death Dis. 6, e1781
(2015).
219. Stambolsky, P. et al. Modulation of the vitamin D3
response by cancer-associated mutant p53. Cancer
Cell 17, 273–285 (2010).
220. Salehi-Tabar, R. et al. Vitamin D receptor as a master
regulator of the c-MYC/MXD1 network. Proc. Natl
Acad. Sci. USA 109, 18827–18832 (2012).
221. Reichrath, J., Saternus, R. & Vogt, T. Endocrine actions
of vitamin D in skin: relevance for photocarcinogenesis of
non-melanoma skin cancer, and beyond. Mol. Cell.
Endocrinol. 453, 96–102 (2017).
222. Stenn, K. S. & Paus, R. Controls of hair follicle cycling.
Physiol. Rev. 81, 449–494 (2001).
223. Morris, R. J. et al. Capturing and profiling adult hair
follicle stem cells. Nat. Biotechnol. 22, 411–417
(2004).
224. Schneider, M. R., Schmidt-Ullrich, R. & Paus, R. The
hair follicle as a dynamic miniorgan. Curr. Biol. 19,
R132–R142 (2009).
225. Gonzales, K. A. U. & Fuchs, E. Skin and its
regenerative powers: an alliance between stem cells
and their niche. Dev. Cell 43, 387–401 (2017).
226. Hochberg, Z. et al. Calcitriol-resistant rickets with
alopecia. Arch. Dermatol. 121, 646–647 (1985).
227. Marx, S. J., Bliziotes, M. M. & Nanes, M. Analysis of
the relation between alopecia and resistance to
1,25-dihydroxyvitamin D. Clin. Endocrinol. 25,
373–381 (1986).
228. Sakai, Y. & Demay, M. B. Evaluation of keratinocyte
proliferation and differentiation in vitamin D receptor
knockout mice. Endocrinology 141, 2043–2049
(2000).
229. Teichert, A. E., Elalieh, H., Elias, P. M., Welsh, J. &
Bikle, D. D. Overexpression of hedgehog signaling is
associated with epidermal tumor formation in vitamin
D receptor-null mice. J. Invest. Dermatol. 131,
2289–2297 (2011).
230. Panteleyev, A. A., Botchkareva, N. V., Sundberg, J. P.,
Christiano, A. M. & Paus, R. The role of the hairless
(hr) gene in the regulation of hair follicle catagen
transformation. Am. J. Pathol. 155, 159–171 (1999).
231. Miller, J. et al. Atrichia caused by mutations in the
vitamin D receptor gene is a phenocopy of generalized
atrichia caused by mutations in the hairless gene.
J. Invest. Dermatol. 117, 612–617 (2001).
232. Ahmad, W. et al. Alopecia universalis associated with
a mutation in the human hairless gene. Science 279,
720–724 (1998).
233. Millar, S. E. Molecular mechanisms regulating hair
follicle development. J. Invest. Dermatol. 118,
216–225 (2002).
234. Xie, Z., Chang, S., Oda, Y. & Bikle, D. D. Hairless
suppresses vitamin D receptor transactivation in
human keratinocytes. Endocrinology 147, 314–323
(2006).
235. Hsieh, J. C. et al. Analysis of hairless corepressor
mutants to characterize molecular cooperation with
the vitamin D receptor in promoting the mammalian
hair cycle. J. Cell. Biochem. 110, 671–686 (2010).
236. Hsieh, J. C. et al. Physical and functional interaction
between the vitamin D receptor and hairless
corepressor, two proteins required for hair cycling.
J. Biol. Chem. 278, 38665–38674 (2003).
237. Cianferotti, L., Cox, M., Skorija, K. & Demay, M. B.
Vitamin D receptor is essential for normal keratinocyte
stem cell function. Proc. Natl Acad. Sci. USA 104,
9428–9433 (2007).
238. Huelsken, J., Vogel, R., Erdmann, B., Cotsarelis, G.
& Birchmeier, W. β-Catenin controls hair follicle
morphogenesis and stem cell differentiation in the
skin. Cell 105, 533–545 (2001).
239. Lisse, T. S. et al. The vitamin D receptor is required
for activation of cWnt and hedgehog signaling in
keratinocytes. Mol. Endocrinol. 28, 1698–1706
(2014).
240. Teichert, A., Elalieh, H., Elias, P., Welsh, J. & Bikle, D.
Over-expression of hedgehog signaling is associated
with epidermal tumor formation in vitamin D receptor
null mice. J. Invest. Dermatol. 131, 2289–2297 (2011).
241. Zarach, J. M., Beaudoin, G. M. 3rd, Coulombe, P. A. &
Thompson, C. C. The co-repressor hairless has a role
in epithelial cell differentiation in the skin.
Development 131, 4189–4200 (2004).
242. DasGupta, R., Rhee, H. & Fuchs, E. A developmental
conundrum: a stabilized form of β-catenin lacking the
transcriptional activation domain triggers features
of hair cell fate in epidermal cells and epidermal cell
fate in hair follicle cells. J. Cell Biol. 158, 331–344
(2002).
243. Daroach, M., Narang, T., Saikia, U. N., Sachdeva, N.
& Sendhil Kumaran, M. Correlation of vitamin D and
vitamin D receptor expression in patients with
alopecia areata: a clinical paradigm. Int. J. Dermatol.
57, 217–222 (2018).
244. Lee, S., Kim, B. J., Lee, C. H. & Lee, W. S. Increased
prevalence of vitamin D deficiency in patients with
alopecia areata: a systematic review and meta-analysis.
J. Eur. Acad. Dermatol. Venereol. 32, 1214–1221
(2018).
245. Umar, M. et al. Vitamin D and the pathophysiology of
inflammatory skin diseases. Skin Pharmacol. Physiol.
31, 74–86 (2018).
246. Mady, L. J. et al. The transient role for calcium and
vitamin D during the developmental hair follicle cycle.
J. Invest. Dermatol. 136, 1337–1345 (2016).
247. Greenlee, R. T., Hill-Harmon, M. B., Murray, T. & Thun, M.
Cancer statistics, 2001. CA Cancer J. Clin. 51, 15–36
(2001).
248. Reichrath, J. et al. Analysis of 1,25-dihydroxyvitamin D(3)
receptors (VDR) in basal cell carcinomas. Am. J. Pathol.
155, 583–589 (1999).
249. Reichrath, J. et al. Analysis of the vitamin D system in
cutaneous squamous cell carcinomas. J. Cutan. Pathol.
31, 224–231 (2004).
250. Ratnam, A. V., Bikle, D. D., Su, M. J. & Pillai, S.
Squamous carcinoma cell lines fail to respond to
1,25-dihydroxyvitamin D despite normal levels
of the vitamin D receptor. J. Invest. Dermatol. 106,
522–525 (1996).
251. Rheinwald, J. G. & Beckett, M. A. Defective terminal
differentiation in culture as a consistent and selectable
character of malignant human keratinocytes. Cell 22,
629–632 (1980).
252. Xie, Z. et al. Phospholipase C-γ1 is required for the
epidermal growth factor receptor-induced squamous
cell carcinoma cell mitogenesis. Biochem. Biophys.
Res. Commun. 397, 296–300 (2010).
253. Zinser, G. M., Sundberg, J. P. & Welsh, J. Vitamin D(3)
receptor ablation sensitizes skin to chemically induced
tumorigenesis. Carcinogenesis 23, 2103–2109 (2002).
254. Ellison, T. I., Smith, M. K., Gilliam, A. C. &
Macdonald, P. N. Inactivation of the vitamin D
receptor enhances susceptibility of murine skin to
UV-induced tumorigenesis. J. Invest. Dermatol. 128,
2508–2517 (2008).
255. Bikle, D. D., Oda, Y., Tu, C. L. & Jiang, Y. Novel
mechanisms for the vitamin D receptor (VDR) in the
skin and in skin cancer. J. Steroid Biochem. Mol. Biol.
148, 47–51 (2015).
256. Tang, J. Y. et al. Vitamin D in cutaneous carcinogenesis:
part I. J. Am. Acad. Dermatol. 67, 803.e1–803.e12
(2012).
257. Tang, J. Y. et al. Vitamin D in cutaneous carcinogenesis:
part II. J. Am. Acad. Dermatol. 67, 817.e1–817.e11
(2012).
258. Bikle, D. D., Jiang, Y., Nguyen, T., Oda, Y. & Tu, C. L.
Disruption of vitamin D and calcium signaling in
keratinocytes predisposes to skin cancer. Front.
Physiol. 7, 296 (2016).
259. Hussein, M. R. Ultraviolet radiation and skin cancer:
molecular mechanisms. J. Cutan. Pathol. 32,
191–205 (2005).
260. Chen, R. H., Maher, V. M. & McCormick, J. J. Effect of
excision repair by diploid human fibroblasts on the
kinds and locations of mutations induced by (+/-)-7
beta,8 alpha-dihydroxy-9 alpha,10 alpha-epoxy-
7,8,9,10- tetrahydrobenzo[a]pyrene in the coding
region of the HPRT gene. Proc. Natl Acad. Sci. USA
87, 8680–8684 (1990).
261. Wood, R. D. DNA damage recognition during
nucleotide excision repair in mammalian cells.
Biochimie 81, 39–44 (1999).
262. Demetriou, S. K. et al. Vitamin D receptor mediates
DNA repair and is UV inducible in intact epidermis but
not in cultured keratinocytes. J. Invest. Dermatol.
132, 2097–2100 (2012).
263. Gupta, R. et al. Photoprotection by 1,25
dihydroxyvitamin D3 is associated with an increase in
p53 and a decrease in nitric oxide products. J. Invest.
Dermatol. 127, 707–715 (2007).
264. Moll, P. R., Sander, V., Frischauf, A. M. & Richter, K.
Expression profiling of vitamin D treated primary
human keratinocytes. J. Cell. Biochem. 100, 574–592
(2007).
265. Ping, X. L. et al. PTCH mutations in squamous cell
carcinoma of the skin. J. Invest. Dermatol. 116,
614–616 (2001).
266. Aszterbaum, M. et al. Identification of mutations in
the human PATCHED gene in sporadic basal cell
carcinomas and in patients with the basal cell nevus
syndrome. J. Invest. Dermatol. 110, 885–888 (1998).
267. Hahn, H. et al. Mutations of the human homolog of
DROSOPHILA patched in the nevoid basal cell
carcinoma syndrome. Cell 85, 841–851 (1996).
268. Regl, G. et al. The zinc-finger transcription factor GLI2
antagonizes contact inhibition and differentiation of
human epidermal cells. Oncogene 23, 1263–1274
(2004).
269. Regl, G. et al. Human GLI2 and GLI1 are part of a
positive feedback mechanism in basal cell carcinoma.
Oncogene 21, 5529–5539 (2002).
270. Grachtchouk, M. et al. Basal cell carcinomas in mice
overexpressing Gli2 in skin. Nat. Genet. 24, 216–217
(2000).
271. Chan, E. F., Gat, U., McNiff, J. M. & Fuchs, E. A common
human skin tumour is caused by activating mutations in
β-catenin. Nat. Genet. 21, 410–413 (1999).
272. Mercer, T. R., Dinger, M. E. & Mattick, J. S. Long
non-coding RNAs: insights into functions. Nat. Rev.
Genet. 10, 155–159 (2009).
www.nature.com/nrendo

273. Mattick, J. S. Long noncoding RNAs in cell and
developmental biology. Semin. Cell Dev. Biol. 22, 327
(2011).
274. Jiang, Y. J. & Bikle, D. D. LncRNA: a new player in 1α,
25(OH)(2) vitamin D(3) /VDR protection against skin
cancer formation. Exp. Dermatol. 23, 147–150
(2014).
275. Underhill, C. CD44: the hyaluronan receptor. J. Cell
Sci. 103, 293–298 (1992).
276. Screaton, G. R. et al. Genomic structure of DNA
encoding the lymphocyte homing receptor CD44
reveals at least 12 alternatively spliced exons. Proc.
Natl Acad. Sci. USA 89, 12160–12164 (1992).
277. Haggerty, J. G., Bretton, R. H. & Milstone, L. M.
Identification and characterization of a cell surface
proteoglycan on keratinocytes. J. Invest. Dermatol.
99, 374–380 (1992).
278. Averbeck, M. et al. Differential regulation of
hyaluronan metabolism in the epidermal and dermal
compartments of human skin by UVB irradiation.
J. Invest. Dermatol. 127, 687–697 (2007).
279. Stern, R. Complicated hyaluronan patterns in skin:
enlightenment by UVB? J. Invest. Dermatol. 127,
512–513 (2007).
280. Bourguignon, L. Y. et al. Selective matrix (hyaluronan)
interaction with CD44 and RhoGTPase signaling
promotes keratinocyte functions and overcomes
age-related epidermal dysfunction. J. Dermatol. Sci.
72, 32–44 (2013).
281. Bourguignon, L. Y. & Bikle, D. Selective hyaluronan–
CD44 signaling promotes miRNA-21 expression and
interacts with vitamin D function during cutaneous
squamous cell carcinomas progression following UV
irradiation. Front. Immunol. 6, 224 (2015).
282. Hsu, J. Y., Feldman, D., McNeal, J. E. & Peehl, D. M.
Reduced 1α-hydroxylase activity in human prostate
cancer cells correlates with decreased susceptibility to
25-hydroxyvitamin D3-induced growth inhibition.
Cancer Res. 61, 2852–2856 (2001).
283. Brożyna, A. A., Jóźwicki, W., Janjetovic, Z. &
Slominski, A. T. Expression of the vitamin D-activating
Nature Reviews | Endocrinology
enzyme 1α-hydroxylase (CYP27B1) decreases during
melanoma progression. Hum. Pathol. 44, 374–387
(2013).
284. Anderson, M. G., Nakane, M., Ruan, X., Kroeger, P. E. &
Wu-Wong, J. R. Expression of VDR and CYP24A1 mRNA
in human tumors. Cancer Chemother. Pharmacol. 57,
234–240 (2006).
285. Solomon, C., White, J. H. & Kremer, R.
Mitogen-activated protein kinase inhibits
1,25-dihydroxyvitamin D3-dependent signal
transduction by phosphorylating human retinoid
X receptor α. J. Clin. Invest. 103, 1729–1735
(1999).
286. Larriba, M. J. et al. Snail2 cooperates with Snail1 in
the repression of vitamin D receptor in colon cancer.
Carcinogenesis 30, 1459–1468 (2009).
287. Mittal, M. K., Myers, J. N., Misra, S., Bailey, C. K.
& Chaudhuri, G. In vivo binding to and functional
repression of the VDR gene promoter by SLUG in
human breast cells. Biochem. Biophys. Res. Commun.
372, 30–34 (2008).
288. Marik, R. et al. DNA methylation-related vitamin D
receptor insensitivity in breast cancer. Cancer Biol.
Ther. 10, 44–53 (2010).
289. Mohri, T., Nakajima, M., Takagi, S., Komagata, S.
& Yokoi, T. MicroRNA regulates human vitamin D
receptor. Int. J. Cancer 125, 1328–1333 (2009).
290. Reichrath, J., Reichrath, S., Heyne, K., Vogt, T. &
Roemer, K. Tumor suppression in skin and other
tissues via cross-talk between vitamin D- and
p53-signaling. Front. Physiol. 5, 166 (2014).
291. Sequeira, V. B. et al. The role of the vitamin D
receptor and ERp57 in photoprotection by 1α,25-
dihydroxyvitamin D3. Mol. Endocrinol. 26, 574–582
(2012).
292. Chen, H. et al. Vitamin D directly regulates Mdm2
gene expression in osteoblasts. Biochem. Biophys.
Res. Commun. 430, 370–374 (2013).
293. Heyne, K., Heil, T. C., Bette, B., Reichrath, J. &
Roemer, K. MDM2 binds and inhibits vitamin D
receptor. Cell Cycle 14, 2003–2010 (2015).
Reviews
294. Dixon, K. M. et al. 1α,25(OH)(2)-vitamin D and a
nongenomic vitamin D analogue inhibit ultraviolet
radiation-induced skin carcinogenesis. Cancer Prev.
Res. 4, 1485–1494 (2011).
295. Skrajnowska, D. & Bobrowska-Korczak, B. Potential
molecular mechanisms of the anti-cancer activity of
vitamin D. Anticancer Res. 39, 3353–3363 (2019).
296. Plikus, M. V. et al. Epithelial stem cells and
implications for wound repair. Semin. Cell Dev. Biol.
23, 946–953 (2012).
297. Oda, Y. et al. Combined deletion of the vitamin D
receptor and calcium-sensing receptor delays wound
re-epithelialization. Endocrinology 158, 1929–1938
(2017).
298. Palmer, H. G., Martinez, D., Carmeliet, G. & Watt, F. M.
The vitamin D receptor is required for mouse hair cycle
progression but not for maintenance of the epidermal
stem cell compartment. J. Invest. Dermatol. 128,
2113–2117 (2008).
299. Luderer, H. F., Nazarian, R. M., Zhu, E. D. & Demay, M. B.
Ligand-dependent actions of the vitamin D receptor
are required for activation of TGF-β signaling during
the inflammatory response to cutaneous injury.
Endocrinology 154, 16–24 (2013).
Author contributions
The authors contributed equally to all aspects of the article.
Competing interests
The authors declare no competing interests.
Peer review information
Nature Reviews Endocrinology thanks M. Holick, G. Jones,
M. Hewison and the other, anonymous, reviewer(s) for their
contribution to the peer review of this work.
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