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Evolution of sexual reproduction: A view from the

fungal kingdom supports an evolutionary epoch
with sex before sexes

Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, NC 27710, USA

article info abstract

Article history: Sexual reproduction is conserved throughout each supergroup within the eukaryotic tree
Received 16 February 2015 of life, and therefore thought to have evolved once and to have been present in the last eu-
Received in revised form karyotic common ancestor (LECA). Given the antiquity of sex, there are features of sexual
10 June 2015 reproduction that are ancient and ancestral, and thus shared in diverse extant organisms.
Accepted 17 August 2015 On the other hand, the vast evolutionary distance that separates any given extant species
from the LECA necessarily implies that other features of sex will be derived. While most
Keywords: types of sex we are familiar with involve two opposite sexes or mating types, recent studies
Evolution in the fungal kingdom have revealed novel and unusual patterns of sexual reproduction,
Heterothallic including unisexual reproduction. In this mode of reproduction a single mating type can
Homothallic on its own undergo self-fertile/homothallic reproduction, either with itself or with other
Inbreeding members of the population of the same mating type. Unisexual reproduction has arisen
MAT independently as a derived feature in several different lineages. That a myriad of different
Mating type types of sex determination and sex determinants abound in animals, plants, protists, and
Meiosis fungi suggests that sex specification itself may not be ancestral and instead may be a
Outcrossing derived trait. If so, then the original form of sexual reproduction may have been unisexual,
Selfing onto which sexes were superimposed as a later feature. In this model, unisexual reproduc-
Sex chromosomes tion is both an ancestral and a derived trait. In this review, we consider what is new and
Sex determination what is old about sexual reproduction from the unique vantage point of the fungal
Sexual reproduction kingdom.
ª 2015 The Author. Published by Elsevier Ltd. This is an open access article under the CC
BY-NC-ND license (

1. Introduction nucleus, mitochondria, secretory apparatus, RNAi, and repro-

ducing both asexually and sexually. Thus, when we think of
No one knows the exact nature of the LECA, but we think that where sex first evolved, it was in the water, involving swim-
this ancestor was a unicellular, aquatic, motile creature with ming cells (Levin and King, 2013; Umen and Heitman, 2013).
one or two flagella. Thus, in some respects the LECA was sim- And when we think of how sex first evolved, this involved
ple. But in other ways, it was already quite complex, with a changes in ploidy and the process of meiosis, given their

* Tel.: þ1 919 684 2824; fax: þ1 919 684 5458.

E-mail address:
1749-4613/ ª 2015 The Author. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://
Evolution of sexual reproduction 109

conserved nature throughout eukaryotes. And while cellecell humans and in mice. But in other plants and animals, there
and nuclearenuclear fusion play prominent roles in sexual are different mechanisms of sex determination. Some species,
reproduction today, there may have been an era in which such as the plant Papaya and the fish Medaka, have sex chro-
endoreplication cycles followed by meiosis drove the pro- mosomes in which the sex specific region is small and the two
cesses of ploidy change during ancestral modes of sexual sex chromosomes are the same size, so called homomorphic
reproduction. In this view, cellecell fusion may be ancient, sex chromosomes (Kondo et al., 2004; Liu et al., 2004; Myosho
but perhaps not as ancient as other features of sexual et al., 2012). Chickens and other birds have a completely
reproduction. different type of sex chromosome, called Z and W, and in
Why sex is so pervasive is thought to result from potential these lineages it is the heterogametic ZW pattern that spec-
benefits conferred by sexual reproduction. These include ifies female and the homogametic ZZ the male (Zhou et al.,
purging the genome of deleterious mutations and shuffling 2014). In some animals, including turtles and crocodiles, the
the genome via independent chromosomal assortment and temperature at which an egg hatches determines sexual iden-
recombination to give rise to a diverse repertoire of meiotic tity and this is called Environmental Sex Determination (ESD)
progeny. Sex may also enable organisms to keep pace with to distinguish it from Chromosomal Sex Determination (CSD)
or outrun pathogens, including those both external and those (Barske and Capel, 2008). Yet other species appear to be hy-
internal (such as transposons). There is sound experimental brids of the two with features of both environmental and
evidence from studies in Caenorhabditis elegans and in natu- chromosomal sex determination. Finally in some lines of the
rally occurring snails in New Zealand for this last hypothesis zebra fish Danio rerio sex appears to be a quantitative trait, in
in which sex allows species to keep pace with their pathogens which genes on multiple different chromosomes come
(King et al., 2009, 2011; Morran et al., 2011; Vergara et al., 2013). together in allelic combinations that favor either female or
However, these potential benefits of sex are pitted against male fate (Anderson et al., 2012; Bradley et al., 2011; Liew
well-known costs of sexual reproduction: that only 50 % of a et al., 2012; Liew and Orban, 2014). This quantitative sex deter-
parental genome is transmitted to any given progeny, the mining system has been termed polygenic sex determination
time and energy required to locate mates, and the breaking (PSD). Recent studies of wild D. rerio reveal a sex determining
apart of well adapted genomic configurations. region on one end of chromosome 4 that may be consistent
The core features of sexual reproduction are conserved in with a WZ/ZZ sex chromosome system, suggesting loss of a
organisms as diverse as the model budding yeast Saccharo- sex determinant or recent origin of a novel one during domes-
myces cerevisiae and humans, despite a billion years or more tication (Wilson et al., 2014). To summarize, in simple terms
of evolution separating us from our last common shared the ways in which sex is determined are plastic and diverse.
ancestor. These conserved features include: 1) ploidy changes What about fungi? Relatively few fungi have large size
from haploid to diploid to haploid (or diploid to haploid to dimorphic sex chromosomes, but there are a few well studied
diploid), 2) the process of meiosis that enables meiotic recom- examples such as Neurospora tetrasperma, Podospora, and Micro-
bination and halves the ploidy of the genome, and 3) cellecell botryum (Ellison et al., 2011; Fraser et al., 2004; Fraser and
fusion between mating partners (a and a cells) or gametes (the Heitman, 2004; Grognet et al., 2014; Hood et al., 2013; Menkis
sperm and the egg). This ubiquity of the conserved features of et al., 2008; Whittle et al., 2015). Most fungi have relatively
sex again speaks to the antiquity of the process. smaller regions of their genome, called mating-type loci, or
Beyond the commonalities in the mechanisms of sex, there MAT for short, that dictate their mating type (Fraser and
are also shared features to the modes of sexual reproduction. Heitman, 2003). The paradigmatic example is S. cerevisiae in
This includes outbreeding between genetically divergent which a relatively small region of the genome, less than a
members of the population, but also types of inbreeding that thousand base pairs, expresses in the alternate mating types
can involve the ability of the yeast S. cerevisiae to undergo mat- one or two key cell fate determinants, all of which are tran-
ing type switching that allows mother cells to mate with their scription factors responsible for orchestrating both haploid
daughter cells. And in humans there are the examples of cell type specificity (a or a) and the diploid zygote fate (a/a).
consanguineous marriages, resulting for example from cous- Two are homeodomain proteins of the HD1 and HD2 class
inecousin pairings, which lead to considerable inbreeding that form a heterodimer, a1/a2, which is necessary for the
with the risk of exposure of recessive alleles in a homozygous diploid zygote fate. The other factor, a1 from MATa, encodes
configuration. We will return to this theme of the balance be- an alpha domain transcription factor necessary for turning
tween outbreeding and inbreeding. on genes required for the a cell fate, while a2 represses a genes
to further enforce the a cell haploid fate. The a haploid cell
type is the default, and is not actively specified by the MAT lo-
2. Mechanisms of sex determination cus. This type of mating-type system is called bipolar to reflect
the two mating types, a and a. When the two mating types are
Sex in humans and many other animals is determined by the in balance in the population, bipolar mating systems enable
X and Y sex chromosomes, in which individuals with XX kar- 50 % outcrossing and 50 % inbreeding.
yotype are female and those with XY are male. The two sex But other fungi have much more exotic sex lives, and have
chromosomes are dramatically different in size, and are a more complex mating-type determining system in which
referred to as heteromorphic sex chromosomes. A single there are literally thousands and thousands and thousands
gene resident on the Y chromosome, SRY, is sufficient to of different mating types (Brown and Casselton, 2001;
direct male fate and transferring this single gene from the Y Casselton, 2002, 2008; Heitman et al., 2007; Raper, 1966). In
to the X chromosome suffices to cause sex reversal in both these species there are two loci that lie unlinked on different
110 J. Heitman

chromosomes that specify mating type. These are called the A Interestingly, there are examples of species with bipolar
and B MAT loci, and one encodes the homeodomain factors, mating type within the Basidiomycota phylum. Are these
and the other encodes pheromones and pheromone receptors remnants of the bipolar ancestral state, like the microwave
(which locus encodes which genes depends on the species, echoes of the Big Bang that formed our universe, or rather
because these loci were named historically as they were derived from the tetrapolar configuration? This question has
discovered genetically long before their molecular basis was been addressed in some detail in the Cryptococcus pathogenic
elucidated). In many species, both loci are multiallelic, and species complex (Heitman et al., 2011). All of the pathogenic
as a result there are many different mating types. Both loci species and lineages, including Cryptococcus neoformans var.
must differ for productive mating and thus an isolate of grubii (serotype A), C. neoformans var. neoformans (serotype D),
A1B1 mating type can mate with an isolate of A2B2 mating and Cryptococcus gattii (VGI, VGII, VGIII, and VGIV) have just
type, but not with isolates that are A1B2 or A2B1. Because two mating types and are bipolar species. Analysis of a series
there are thousands of different mating types, most encoun- of closely aligned and related species reveals that multiple
ters between isolates in nature will be fertile, and this drives outgroup species have tetrapolar mating systems. This in-
the frequency of outbreeding to >99 %. On the other hand, cludes Cryptococcus amylolentus and Cryptococcus heveanensis,
from any given cross, say A1B1 by A2B2, because two MAT for which our group discovered extant sexual cycles, cloned
loci segregate independently four different types of progeny and characterized their mating-type loci, and showed defini-
are produced (A1B1, A2B2, A1B2, and A2B1) and these systems tively that both are tetrapolar (Findley et al., 2012; Metin
are therefore called tetrapolar mating systems. Because any et al., 2010). Related studies for Kwoniella mangrovensis charac-
given progeny is only interfertile with 25 % of its siblings terized its mating type locus and clarified that it is tetrapolar
(A1B2 can mate with A2B1 but not with A1B1 or A2B2), the tet- (Guerreiro et al., 2013), in contrast to a previous report from
rapolar configuration not only promotes outcrossing but also others that had concluded it was bipolar (Statzell-Tallman
leads to inbreeding depression. It is thought to be these differ- et al., 2008). Taken from this vantage point of multiple tetrapo-
ences in the frequency of outcrossing and inbreeding that pro- lar outgroup closely aligned species, we can conclude that the
vided the evolutionary pressure for transitions between bipolar configuration of the pathogenic species complex is a
bipolar and tetrapolar mating systems. derived state and has a monophyletic origin. E pluribus duo!
Phylogenetic reconstructions across the fungal kingdom The longer evolutionary pathway was thus bipolar to tetrapo-
support the conclusion that bipolar mating type is an ances- lar and then a return to bipolar. In this view then the bipolar
tral state and the tetrapolar configuration is a derived state. state is both an ancestral state and a derived one.
If we examine species in the Ascomycota and Zygomycota/ The transition from tetrapolar to bipolar has occurred
Mucorales, all species with known mating type systems are bi- several times independently in the Basidiomycota, and other
polar. This includes a myriad of species throughout the Asco- examples include Ustilago hordei and Malassezia species
mycota including S. cerevisiae (a and a), Schizosaccharomyces (Bakkeren et al., 2006; Bakkeren and Kronstad, 1994; Gioti
pombe (P and M), Candida albicans (a/a and a/a), Neurospora et al., 2013; Hsueh and Heitman, 2008; Lee et al., 1999). That
crassa (A and a), and Aspergillus fumigatus (MAT1-1 and the cases that are known involve pathogens of either humans
MAT1-2) just to name a few (Glass et al., 1988; Hull et al., or plants suggests there may be a causal association between
2000; Magee and Magee, 2000; O’Gorman et al., 2009). In the restricted outbreeding and enhanced inbreeding and patho-
Mucorales, we again find just two mating types, typically genesis. Perhaps as microbial pathogens become well adapted
called P and M for plus (þ) and minus (), in Phycomyces blake- to their hosts, less rather than more genetic exchange is adap-
sleeanus, Mucor circinelloides, and Rhizopus oryzae (Gryganskyi tive. They clearly do conserve the ability to undergo sex, and
et al., 2010; Idnurm et al., 2008; Lee et al., 2008). This is in thus this may ensure their longer term evolutionary success,
marked contrast to the Basidiomycota branch of the dikarya but restricting genetic exchange may allow them to vary just
in which a majority of species have the tetrapolar mating sys- enough to keep pace with their hosts.
tem configuration. But thus far, no species with the tetrapolar We return then to a more general consideration of MAT loci
mating system have been found outside of the Basidiomycota. and several recent poignant examples prove illuminating.
Thus, we can root the phylogenetic tree with multiple out- Recent studies have defined the MAT loci for two ciliates in
groups that are all bipolar, and conclude that the tetrapolar the Alveolate supergroup (Tetrahymena and Paramecium), for
system is a derived state, possibly with a single origin at the two representative algae in the Planta supergroup (Chlamydo-
base of the Basidiomycota. In essence, an ancestral system monas and Volvox), and for the slime mold Dictyostelium in
with just one MAT locus encoding homeodomain factors the Amoebozoa supergroup (Bloomfield et al., 2010;
evolved into a system with a second sex determinant on Cervantes et al., 2013; Ferris et al., 2010; Singh et al., 2014).
another chromosome encoding pheromones and pheromone While there are fascinating features, such as the three mating
receptors. How such a transition might have occurred could types of the slime mold, and the dramatic expansion of MAT
have first involved an auto-stimulated pheromone, phero- loci into true sex chromosomes from Chlamydomonas to
mone receptor gene pair, which underwent genetic drift to Volvox, and the roulette type mating-type switching of Tetra-
form more than one self-activating allele, followed by recom- hymena which has seven mating types (one for each day of
bination between the two to generate two self-sterile/cross- the week, or seven brides for seven brothers), there is essen-
fertile gene combinations that became a second MAT locus. tially no underlying conserved feature. Taken from the
Some aspects of this have been modeled, conceptualizing perspective of diverse and plastic sex chromosomes and
how such an event might have transpired (Fraser et al., 2007). mating-type loci from at least 5 of the 8 supergroups of the
Evolution of sexual reproduction 111

eukaryotes, there seems to be nothing ancestral or static features associated with bisexual reproduction, with the
about sex determination. exception that it dispenses with the obligate need for a part-
These considerations lead us to conclude that mating type ner of opposite mating type. There are situations, so called
and sex determination are not ancestral, but rather derived me nage a  trois matings, in which three partners are present
features. If so, then what was the ancestral pattern of sexual (two a and one a) and a small number of limiting a cells serve
reproduction? Put another way, we know that the LECA was as pheromone donors to stimulate the fusion and unisexual
sexy but how sexuality was manifested is the central ques- reproduction of two a partners present as the majority in the
tion. As we will consider in the section that follows, our cen- population. In fact in mixed populations, both a-a bisexual
tral premise is that LECA was unisexual (Fig. 1). and aea unisexual reproduction can occur concomitantly
and the balance depends upon the ratio of the mating types
and who encounters whom.
3. Modes of sexual reproduction While early reports focused on unisexual reproduction/
monokaryotic fruiting of isolates of a mating type (Wickes
We now turn our considerations from mechanisms of sexual et al., 1996), we now appreciate that isolates of a mating type
reproduction to modes of sexual reproduction. Consideration can also undergo unisexual reproduction (Hull and Heitman,
of the sexual nature of the Cryptococcus pathogenic species 2002; Tscharke et al., 2003). Subsequently, by detailed genetic
complex provides further illumination. We have discussed analysis of progeny from a cross of a more and a less fertile
how they have undergone a transition from an enhanced out- parental strain, it was found that unisexual reproduction is
crossing/restricted inbreeding ancestral tetrapolar state with a quantitative trait, to which multiple quantitative trait loci
thousands of mating types to a derived bipolar state with (QTL) scattered around the genome contribute (Lin et al.,
just two mating types with enhanced inbreeding and 2006). The MAT locus was found to be the most significant of
restricted outbreeding. We will now consider how they have the QTLs mapped, with the a allele promoting unisexual hy-
taken this transition one step further from a bipolar to a uni- phal development to a greater extent than the a allele (Lin
polar sexual state with just one mating type that reproduces et al., 2006). To put this another way, isolates with the a allele
unisexually, dispensing with the need for a partner of oppo- are skewed towards being of higher fecundity compared to
site mating type. E pluribus unum! isolates with the a allele, but if the other QTLs are those that
A central conundrum in the field was that while there was promote unisexual reproduction they can elicit development
a defined bipolar mating-type system with two mating types in an a mating type background. Why a isolates predominate
that could occur under lab conditions (Kwon-Chung, 1976a), in nature is not known, and could result from a bottleneck
the vast majority of clinical and environmental isolates were when C. neoformans emerged from Africa to become global-
all of just one mating type, a. In many populations no isolates ized, or may result from enhanced fitness under conditions
of a mating type could be identified. This led to the conclusion that have not yet been found.
that the organism might be largely clonal and asexual. Similar Population genetic evidence also supports that unisexual
reasoning was applied to the vast majority of fungal patho- reproduction may be a predominant mode of sexual reproduc-
gens and eukaryotic parasite pathogens of humans, and as tion in nature. This includes detailed analyses of aADa hybrids
recently as a decade ago it was thought that the majority produced via unisex (Lin et al., 2007), and also aAAa diploids
were asexual and clonal. We now appreciate that they are sex- that may be intermediates or products of the unisexual cycle
ual, but in unusual ways, including unisexual and parasexual (Lin et al., 2009). Finally a series of studies have revealed evi-
(Heitman, 2006, 2010). dence of recombination, even in populations that are exclu-
The discovery of unisexual reproduction of Cryptococcus sively of one mating type (Bui et al., 2008; Hiremath et al.,
came from revisiting older observations (Erke, 1976; Wickes 2008; Saul et al., 2008).
et al., 1996) about the production of hyphae, basidia, and The analysis of aAAa diploids reveals two distinct types.
spores by certain isolates cultured solo on a variety of media One class is clearly the progeny of two genetically different
that support mating (V8, MS, SLAD, FA). Several lines of evi- parents, and thus appears to have been generated via aea
dence converged to show that this developmental process, cellecell fusion. The second class has two seemingly identical
which had been called haploid or monokaryotic fruiting and genomes, and thus may have arisen from either endoreplica-
was thought to be strictly asexual, is in fact an unusual form tion or cellecell fusion between a mother and her daughter.
of self-fertile homothallic sexual reproduction (Feretzaki and Our group is currently identifying and studying genes
Heitman, 2013a, b; Lin et al., 2005). The process involves ploidy involved in cellecell fusion and nuclear fusion to test these
changes which, in some cases, can involve cellecell fusion, and other models.
but likely also occur via endoreplication. Key meiotic genes, The fact that some unisexual reproduction appears to
including SPO11 and DMC1, which encode the proteins that involve two genetically identical genomes or isolates chal-
make and repair the DSB DNA breaks that provoke meiotic lenges conventional models for the evolution and impact of
recombination, are required for this developmental sexual sexual reproduction. Put another way, if there is no pre-
process. Our group has recently reported a detailed analysis existing genetic diversity to admix, why go to the trouble of
of meiotic recombination features occurring during bisexual expending so much energy undergoing sexual reproduction?
and unisexual reproduction, further underscoring that core Are these just teenagers spinning their wheels in a parking
features of sexual reproduction occur during both modes of lot, or is there some potential benefit? It is important to
sexual reproduction (Sun et al., 2014). Thus, under laboratory consider that the null hypothesis may be that this mode of
conditions unisexual reproduction has all of the hallmark sexual reproduction is not of benefit, rather that it may have
112 J. Heitman

Fig. 1 e Unisexual reproduction as an ancestral and derived trait. A phylogenetic depiction of the major supergroups of the
Eukaryotic tree of life depicting the LECA as sexual/unisexual in the ancestral state, with a diverse pattern of derived sexual
states including predominantly bisexual extant species but also those that are unisexual, trisexual, septasexual, and mul-
tisexual. Modified from Baldauf (2003) with permission of the author.

little cost and therefore is just tolerated (Lynch, 2007). But this C. albicans has the capacity to undergo both bisexual mating
said, our group has been exploring experimentally if and how and unisexual mating as a prelude to its parasexual cycle
it might confer some type of evolutionary benefit, and have was an important advance (Alby et al., 2009). Thus, we now
published three studies thus far supporting the contention know that two of the most common systemic human fungal
that it may. pathogens have extant sexual/parasexual cycles and the ca-
First, our group has presented evidence that unisexual pacity for unisexual reproduction. Interestingly, there are a
reproduction allows a transition from yeast to hyphae and group of four Neurospora species that have extant homothallic
thereby promotes more efficient foraging for nutrients and (self-fertile) sexual cycles but only one mating type, and thus
also the generation of spores that can be disseminated to likely another independent origin and example of unisexual
more distant locales by wind currents (Phadke et al., 2013). reproduction (Gioti et al., 2012; Glass et al., 1990; Glass and
Second, our research has shown that unisexual reproduction Smith, 1994). Similarly, a group of Stemphylium isolates repre-
can generate genetic diversity de novo, and that much of this senting a novel self-fertile group of isolates with only the
diversity results from aneuploidy as a consequence of MAT1-1 mating type information at the MAT locus and no iso-
meiosis, and that this occurs during both unisexual and lates with the MAT1-2 information represents another inde-
bisexual reproduction (Ni et al., 2013). Third, our studies pendent example and origin of unisexual reproduction
recently reported indicate that unisex has the capacity to (Inderbitzin et al., 2005). A very recent study reports that Pneu-
turn back the hands of time and reverse Muller’s ratchet and mocystis species, which are obligate fungal pathogens that
prevent the otherwise inexorable accumulation of deleterious infect and only live in animal lungs, have a fusion of the two
mutations (Roach and Heitman, 2014), the process that is opposite mating types related to the S. pombe MAT loci, sug-
thought to be that which dooms asexual species to inevitable gesting that Pneumocystis is homothallic/self-fertile and may
extinction. There are a variety of possible benefits that are in be undergoing its sexual cycle in the lungs of infected animals
the process of being tested experimentally, some of which (Almeida et al., 2015). This is not strictly a case of unisexual
have been reviewed (Roach et al., 2014). reproduction because both MAT loci are present in a fused
Things that are found once in biology are interesting, but configuration rather than just one MAT locus, but it does serve
those that are found more than once are more interesting to further illustrate how commonly pathogenic microbes have
because they may be generalizable. Thus, the discovery that undergone the transition from outcrossing heterothallic
Evolution of sexual reproduction 113

sexual reproduction to homothallic self-fertile inbreeding. It is for recombination in the population. Despite prodigious effort,
important to note that these types of fused mating-type loci no extant sexual cycle has as yet been found, but it seems
leading to self-fertile homothallic fungal species had been likely that some clever genetic trick, or specific environmental
described more than a decade earlier by Gillian Turgeon and condition, or combination of fertile strains, remains to be
colleagues in now classic studies of plant fungal pathogens, discovered.
including Cochliobolus sp. and Gibberella sp., illustrating If a fungus were asexual, how would we recognize it? It is
convergent evolution to homothallism in diverse fungal line- often posited that when a pathway is lost that the genes asso-
ages (Yun et al., 2000, 1999). ciated with it rapidly decay into pseudogenes and then are
This review is dedicated to considering sexual reproduc- lost. And yet recent studies from Greg Lang and David Botstein
tion in the fungal kingdom, so we will just note in passing reveal that sterile variants of S. cerevisiae rapidly arise during
that it is also the case that multiple eukaryotic parasitic path- asexual vegetative growth (Lang et al., 2011, 2009). They
ogens including Toxoplasma, Giardia, Trypanosoma, and Leish- discovered that 23 genes activated by the mating pheromone
mania appear to undergo selfing forms of sexual response pathway are expressed at some level even during
reproduction (Heitman, 2010; Poxleitner et al., 2008; Wendte asexual growth. Mutants that inactivate any step in the pher-
et al., 2010). Nothing is as yet known about whether they omone response pathway grow w2 % faster than their compa-
have mating types and if so how these are configured, but it triots because they no longer express these 23 messages and
does serve to illustrate common features shared between the encoded proteins. And relieved of this gene expression
fungal and parasite eukaryotic pathogens. deadweight, they are more fit as asexual competitors.
I wish to return then to our question of what the sexuality Sequence of their genomes reveals a variety of simple muta-
of the LECA ancestor of all eukaryotes looked like and propose tions in many steps in the pheromone response pathway.
that there was an evolutionary epoch in which there was sex But the pathway has not yet had time to decay away, and
before sexes and in which unisexual reproduction was the once the first mutation has occurred to relieve the burden of
mode of sexual reproduction. In this view, sexes and mating futile gene expression, there is very little additional benefit
types were then added later, superimposed on an earlier conferred by loss of other genes in the pathway. Thus after a
ancestral system that was unisexual. In this model, the uni- very quick coup de gra ^ ce, the decline of the rest of the
sexual state is then both an ancestral state and a derived pathway is likely to be anything but swift. Put another way,
one. And in this view, the pathogenic microbes have not the rate of decay of the pathway is not constant, and there
invented an entirely new mode of sexual reproduction out of is a very rapid first step followed by a series of much slower
whole cloth. Rather they have returned to an ancestral model steps. This is interesting for several reasons. First, it appears
of reproduction in an evolutionary pathway involving transi- to be very easy and even beneficial to evolve asexuality, at
tions from unisexual to bisexual and back again to unisexual. least for some clones in a population. Second, genome inspec-
It seems fitting to close this section with a quote: tion of an unknown species may lull us into complacency
when we find the mating and meiotic genes seem to be largely
“We shall not cease from exploration intact; it seems likely that a single nucleotide variant (SNV) in
one gene in either pathway might be easily missed. Finally,
And the end of all our exploring evolutionary paths are often thought to be unidirectional.
Losing sex and becoming asexual is thought to be irreversible.
Will be to arrive where we started
But if the loss of sex is due to one SNV in one gene, this is likely
to be fully reversible given sufficient selective pressure for
And know the place for the first time.”
mating or recombination.
T.S. Eliot, from “Little Gidding”, Four Quartets Other ways in which an asexual fungal species might man-
ifest could involve gross rearrangements of the genome. Reg-
ular repeated rounds of meiosis serve to cull from the
4. What questions remain to be addressed by population those members whose genomes have strayed
studies of fungal sex? from the canonical karyotype. But if a species were to be
asexual, one might find a highly variable karyotype from
I Asexuality? isolate to isolate in the population. And examples of just this
pattern have been observed in Verticillium species and sug-
Given that there are an estimated 5 million species of fungi, gested to reflect asexuality (de Jonge et al., 2013).
and maybe more, much remains to be learned about their sex- What other asexual hallmarks might we search for? It has
ual nature (Blackwell 2011). Many fungi are currently classified been hypothesized that to survive a transition to asexuality,
as asexual, and yet many fungi long thought to be asexual species might need to have shed their entire transposon
have been subsequently found to be sexual, or in some cases load in order to survive. So searching for fungal species whose
at least parasexual (i.e. C. albicans). Are there any truly asexual genomes lack transposons might be a starting place.
fungi? Candida glabrata is one potential candidate, but also a Alternatively, we might explore for fungi that lack one or
mystery in that all known isolates are haploids of one or the more key meiotic genes, such as SPO11 or other genes in the
other of two opposite mating types and much of the machin- meiotic toolkit (Schurko and Logsdon, 2008). Thus far, the
ery associated with mating and meiosis are present in the only known examples of eukaryotes lacking SPO11 are the Dic-
genome (Muller et al., 2008; Ramirez-Zavaleta et al., 2010; Sri- tyostelium species. In those cases, meiosis and meiotic recom-
kantha et al., 2003; Wong et al., 2003). There is even evidence bination are known to occur and it seems plausible and even
114 J. Heitman

likely that some other enzyme has stepped into the shoes of may have been a need to outrun pathogens, either exogenous
SPO11 and serves to produce the lesions that provoke meiotic (bacteria, viruses, parasites, fungi, prions) or endogenous ones
recombination (Goodenough and Heitman, 2014). (transposons).
Another way to frame this key evolutionary transition and
II Why two sexes? question about the facultative to obligately sexual state is to
consider a simple question? Are there any fungi who are obli-
Why are there most commonly two sexes? We do not gately sexual? One might make the case that some fungi such
know the answer to this seemingly simple question, but hy- as basidiomycetous mushrooms spend a significant fraction
potheses abound (Haag, 2007; Schaffer 2007). 1) It may be to of their lifecycle as a dikaryotic mycelium and fruiting bodies,
increase the efficiency of mate recognition and fusion. 2) It and these might be examples of either very frequently sexual
may serve to restrict outbreeding compared to populations or even obligately sexual fungi.
or species in which there is one universal mating type that One interesting species that our group has been studying
can mate with anyone. 3) Having two sexes may be also to as one of the outgroups to the Cryptococcus pathogenic species
restrict inbreeding compared to one universal mater that complex is Filobasidiella depauperata (Rodriguez-Carres et al.,
can mate with itself as well as with anyone else. 4) Having 2010). It is one of three species that are most closely aligned
two mating types or sexes may have evolved as a response with the pathogenic species (Findley et al., 2009), but it itself
to conflicts between nuclear and mitochondrial genomes, or is not a pathogen of animals, but it may be a mycoparasite
between mitochondrial genomes, and thereby given rise to (a parasite of other fungi). This is a particularly fascinating
uniparental inheritance of mitochondria and chloroplasts. species. It grows remarkably slowly, is easily contaminated,
5) Having two defined mating types or sexes may have and is quite heat intolerant and must be cultured at tempera-
enabled specialization of gametes and differential contribu- tures of 24  C or lower. But what is fascinating is that this spe-
tion of resources, giving rise to certain development out- cies has no yeast form, and it is strictly hyphal. Microscopic
comes that we observe such as the transition from examination of its hyphae reveals that it shares features
isogametic species to anisogametic ones. 6) Two sexes might with the hyphae produced during the sexual cycle of Crypto-
have served to enable the development of sex specific and coccus, and the hyphae are decorated with basidia and abun-
sex antagonistic traits. 7) Finally, having two mating types dant, long, glorious chains of spores. Thus, it appears that F.
or sexes may serve to enable organisms to know when they depauperata may be a close relative of Cryptococcus that has
are diploid, as they will be heterozygous rather than homozy- become locked in a permanent sexual state. If so, it may be
gous for sex determinants contributed by each haploid a case of an obligately sexual fungus. That there are only
parent. These possible functions are also not mutually exclu- two isolates known, and their growth is limited and restricted
sive, either with each other or with other possible functions under lab conditions, does not provide a ready explanation for
not considered here. why an obligate sexual state would be more beneficial than a
In the context of considering why there are most facultative one, but given further study, explanations may
commonly two sexes, it is worth remembering that there are emerge. It is at least one vantage point from which to begin
examples in which there are more than two mating types or to address this question.
sexes. These include the basidiomycete fungi we have dis- Coda Some things are new, and others old, and sometimes
cussed that have literally thousands and thousands and thou- what is new is also old.
sands of different mating types (multisexual) including
Coprinopsis cinerea, the slime mold Dictyostelium with its three
mating types (trisexual), and the ciliate Tetrahymena with
seven (septasexual) (see Fig. 1). These may seem exceptions, Acknowledgments
but considering what is shared and what is distinct between
closely aligned species with two sexes vs. those with more, Our research on fungal mating-type loci is supported by NIH/
or those with only one (unisexual, to mean “uni” as in one NIAID R01 grant AI50113-11, and our studies on fungal unisex-
sex, or “universal” as in pansexual and capable of mating ual reproduction are supported by NIH/NIAID R37 MERIT
with any other member of the population as well as itself), award AI39115-18. I thank Soo Chan Lee, Ci Fu, Kevin Roach,
may prove enlightening. and Sheng Sun for comments and suggestions. This review
is dedicated to Lorna Casselton for inspiration and for always
III Why obligate sexuality? asking questions that forced one to think, and to think deeply
and with clarity and conviction.
Many animals are obligately sexual. Why? This is in
marked and striking contrast to the vast majority of microbial
eukaryotes which are facultatively sexual, and happily
capable of vegetative mitotic clonal reproduction interspersed
with bouts of sexual reproduction. Why this is not so for
Alby, K., Schaefer, D., Bennett, R.J., 2009. Homothallic and het-
humans, mice, rabbits, is an interesting conundrum. At the
erothallic mating in the opportunistic pathogen Candida albi-
root it suggests that recombinant progeny may be always bet-
cans. Nature 460, 890e893.
ter than the parental. If so, what selective force might have Almeida, J.M., Cisse, O.H., Fonseca, A., Pagni, M., Hauser, P.M.,
driven the transition from a facultatively sexual last common 2015. Comparative genomics suggests primary homothallism
ancestor to a derived obligately sexual state? One driving force of pneumocystis species. mBio 6 e02250e02214.
Evolution of sexual reproduction 115

Anderson, J.L., Rodriguez Mari, A., Braasch, I., Amores, A., Findley, K., Sun, S., Fraser, J.A., Hsueh, Y.P., Averette, A.F., Li, W.,
Hohenlohe, P., Batzel, P., Postlethwait, J.H., 2012. Multiple sex- Dietrich, F.S., Heitman, J., 2012. Discovery of a modified tet-
associated regions and a putative sex chromosome in zebra- rapolar sexual cycle in Cryptococcus amylolentus and the evo-
fish revealed by RAD mapping and population genomics. PLoS lution of MAT in the Cryptococcus species complex. PLoS Genet.
One 7, e40701. 8, e1002528.
Bakkeren, G., Jiang, G., Warren, R.L., Butterfield, Y., Shin, H., Fraser, J.A., Heitman, J., 2003. Fungal mating-type loci. Curr. Biol.
Chiu, R., Linning, R., Schein, J., Lee, N., Hu, G., Kupfer, D.M., 13, R792eR795.
Tang, Y., Roe, B.A., Jones, S., Marra, M., Kronstad, J.W., 2006. Fraser, J.A., Diezmann, S., Subaran, R.L., Allen, A., Lengeler, K.B.,
Mating factor linkage and genome evolution in basidiomyce- Dietrich, F.S., Heitman, J., 2004. Convergent evolution of
tous pathogens of cereals. Fungal Genet. Biol. 43, 655e666. chromosomal sex-determining regions in the animal and
Bakkeren, G., Kronstad, J.W., 1994. Linkage of mating-type loci fungal kingdoms. PLoS Biol. 2, e384.
distinguishes bipolar from tetrapolar mating in basidiomyce- Fraser, J.A., Heitman, J., 2004. Evolution of fungal sex chromo-
tous smut fungi. Proc. Natl. Acad. Sci. U. S. A. 91, 7085e7089. somes. Mol. Microbiol. 51, 299e306.
Baldauf, S.L., 2003. The deep roots of eukaryotes. Science 300, Fraser, J.A., Hsueh, Y.P., Findley, K., Heitman, J., 2007. Evolution of
1703e1706. the mating-type locus: the basidiomycetes. In: Heitman, J.,
Barske, L.A., Capel, B., 2008. Blurring the edges in vertebrate sex Kronstad, J., Taylor, J.W., Casselton, L.A. (Eds), Sex in Fungi:
determination. Curr. Opin. Genet. Dev. 18, 499e505. Molecular Determination and Evolutionary Implications. ASM
Blackwell, M., 2011. The fungi: 1,2,3.5.1 million species? Am. J. Press, Washington, D.C., pp. 19e34
Bot. 98, 426e438. Gioti, A., Mushegian, A.A., Strandberg, R., Stajich, J.E.,
Bloomfield, G., Skelton, J., Ivens, A., Tanaka, Y., Kay, R.R., 2010. Johannesson, H., 2012. Unidirectional evolutionary transitions
Sex determination in the social amoeba Dictyostelium discoi- in fungal mating systems and the role of transposable ele-
deum. Science 330, 1533e1536. ments. Mol. Biol. Evol. 29, 3215e3226.
Bradley, K.M., Breyer, J.P., Melville, D.B., Broman, K.W., Gioti, A., Nystedt, B., Li, W., Xu, J., Andersson, A., Averette, A.F.,
Knapik, E.W., Smith, J.R., 2011. An SNP-based linkage map for Munch, K., Wang, X., Kappauf, C., Kingsbury, J.M., Kraak, B.,
zebrafish reveals sex determination loci. G3 (Bethesda) 1, 3e9. Walker, L.A., Johansson, H.J., Holm, T., Lehtio, J., Stajich, J.E.,
Brown, A.J., Casselton, L.A., 2001. Mating in mushrooms: Mieczkowski, P., Kahmann, R., Kennell, J.C., Cardenas, M.E.,
increasing the chances but prolonging the affair. Trends Lundeberg, J., Saunders, C.W., Boekhout, T., Dawson, T.L.,
Genet. 17, 393e400. Munro, C.A., de Groot, P.W., Butler, G., Heitman, J.,
Bui, T., Lin, X., Malik, R., Heitman, J., Carter, D., 2008. Isolates of Scheynius, A., 2013. Genomic insights into the atopic eczema-
Cryptococcus neoformans from infected animals reveal genetic associated skin commensal yeast Malassezia sympodialis. mBio
exchange in unisexual, alpha mating type populations. Eu- 4 e00572e00512.
karyot. Cell 7, 1771e1780. Glass, N.L., Vollmer, S.J., Staben, C., Grotelueschen, J.,
Casselton, L.A., 2002. Mate recognition in fungi. Heredity 88, Metzenberg, R.L., Yanofsky, C., 1988. DNAs of the two mating-
142e147. type alleles of Neurospora crassa are highly dissimilar. Science
Casselton, L.A., 2008. Fungal sex genes-searching for the ances- 241, 570e573.
tors. Bioessays 30, 711e714. Glass, N.L., Metzenberg, R.L., Raju, N.B., 1990. Homothallic Sor-
Cervantes, M.D., Hamilton, E.P., Xiong, J., Lawson, M.J., Yuan, D., dariaceae from nature: The absence of strains containing only
Hadjithomas, M., Miao, W., Orias, E., 2013. Selecting one of the a mating type sequence. Exp. Mycol. 14, 274e289.
several mating types through gene segment joining and Glass, N.L., Smith, M.L., 1994. Structure and function of a mating-
deletion in Tetrahymena thermophila. PLoS Biol. 11, e1001518. type gene from the homothallic species Neurospora africana.
de Jonge, R., Bolton, M.D., Kombrink, A., van den Berg, G.C., Mol. Gen. Genet. 244, 401e409.
Yadeta, K.A., Thomma, B.P., 2013. Extensive chromosomal Goodenough, U., Heitman, J., 2014. Origins of eukaryotic sexual
reshuffling drives evolution of virulence in an asexual path- reproduction. Cold Spring Harb. Perspect. Biol. 6 a016154.
ogen. Genome Res. 23, 1271e1282. Grognet, P., Bidard, F., Kuchly, C., Tong, L.C., Coppin, E.,
Ellison, C.E., Stajich, J.E., Jacobson, D.J., Natvig, D.O., Lapidus, A., Benkhali, J.A., Couloux, A., Wincker, P., Debuchy, R., Silar, P.,
Foster, B., Aerts, A., Riley, R., Lindquist, E.A., Grigoriev, I.V., 2014. Maintaining two mating types: structure of the mating
Taylor, J.W., 2011. Massive changes in genome architecture type locus and its role in heterokaryosis in Podospora anserina.
accompany the transition to self-fertility in the filamentous Genetics 197, 421e432.
fungus Neurospora tetrasperma. Genetics 189, 55e69. Gryganskyi, A.P., Lee, S.C., Litvintseva, A.P., Smith, M.E.,
Erke, K.H., 1976. Light microscopy of basidia, basidiospores, and Bonito, G.M., Porter, T.M., Anishchenko, I.M., Heitman, J.,
nuclei in spores and hyphae of Filobasidiella neoformans (Cryp- Vilgalys, R., 2010. Structure, function, and phylogeny of the
tococcus neoformans). J. Bacteriol. 128, 445e455. mating locus in the Rhizopus oryzae complex. PLoS One 5,
Feretzaki, M., Heitman, J., 2013a. Genetic circuits that govern 1e12.
bisexual and unisexual reproduction in Cryptococcus neofor- Guerreiro, M.A., Springer, D.J., Rodrigues, J.A., Rusche, L.N.,
mans. PLoS Genet. 9, e1003688. Findley, K., Heitman, J., Fonseca, A., 2013. Molecular and ge-
Feretzaki, M., Heitman, J., 2013b. Unisexual reproduction drives netic evidence for a tetrapolar mating system in the basidio-
evolution of eukaryotic microbial pathogens. PLoS Pathog. 9, mycetous yeast Kwoniella mangrovensis and two novel sibling
e1003674. species. Eukaryot. Cell 12, 746e760.
Ferris, P., Olson, B.J., De Hoff, P.L., Douglass, S., Casero, D., Haag, E.S., 2007. Why two sexes? Sex determination in multicel-
Prochnik, S., Geng, S., Rai, R., Grimwood, J., Schmutz, J., lular organisms and protistan mating types. Semin. Cell Dev.
Nishii, I., Hamaji, T., Nozaki, H., Pellegrini, M., Umen, J.G., Biol. 18, 348e349.
2010. Evolution of an expanded sex-determining locus in Heitman, J., 2006. Sexual reproduction and the evolution of mi-
Volvox. Science 328, 351e354. crobial pathogens. Curr. Biol. 16, R711eR725.
Findley, K., Rodriguez-Carres, M., Metin, B., Kroiss, J., Fonseca, A., Heitman, J., Kronstad, J.W., Taylor, J.W., Casselton, L.A., 2007. Sex
Vilgalys, R., Heitman, J., 2009. Phylogeny and phenotypic in Fungi: Molecular Determination and Evolutionary Implica-
characterization of pathogenic Cryptococcus species and tions. ASM Press, Washington, D.C.
closely related saprobic taxa in the Tremellales. Eukaryot. Cell Heitman, J., 2010. Evolution of eukaryotic microbial pathogens via
8, 353e361. covert sexual reproduction. Cell Host Microbe 8, 86e99.
116 J. Heitman

Heitman, J., Hsueh, Y.P., Metin, B., Findley, K., Rodriguez- neoformans: evidence of same-sex mating in nature and hybrid
Carres, M., 2011. The mating-type locus of Cryptococcus: evo- fitness. PLoS Genet. 3, 1975e1990.
lution of gene clusters governing sex determination and sex- Lin, X., Patel, S., Litvintseva, A.P., Floyd, A., Mitchell, T.G.,
ual reproduction from the phylogenomic perspective. In: Heitman, J., 2009. Diploids in the Cryptococcus neoformans
Heitman, J., Kozel, T.R., Kwon-Chung, K.J., Perfect, J.R., serotype A population homozygous for the alpha mating type
Casadevall, A. (Eds), Cryptococcus: From Human Pathogen to originate via unisexual mating. PLoS Pathog. 5, e1000283.
Model Yeast. ASM Press, Washington, D.C., pp. 139e149 Liu, Z., Moore, P.H., Ma, H., Ackerman, C.M., Ragiba, M., Yu, Q.,
Hiremath, S.S., Chowdhary, A., Kowshik, T., Randhawa, H.S., Pearl, H.M., Kim, M.S., Charlton, J.W., Stiles, J.I., Zee, F.T.,
Sun, S., Xu, J., 2008. Long-distance dispersal and recombina- Paterson, A.H., Ming, R., 2004. A primitive Y chromosome in
tion in environmental populations of Cryptococcus neoformans papaya marks incipient sex chromosome evolution. Nature
var. grubii from India. Microbiology 154, 1513e1524. 427, 348e352.
Hood, M.E., Petit, E., Giraud, T., 2013. Extensive divergence be- Lynch, M., 2007. The frailty of adaptive hypotheses for the origins
tween mating-type chromosomes of the anther-smut fungus. of organismal complexity. Proc. Natl. Acad. Sci. U. S. A. 104
Genetics 193, 309e315. (Suppl 1), 8597e8604.
Hsueh, Y.-P., Heitman, J., 2008. Orchestration of sexual repro- Magee, B.B., Magee, P.T., 2000. Induction of mating in Candida al-
duction and virulence by the fungal mating-type locus. Curr. bicans by construction of MTLa and MTLa strains. Science 289,
Opin. Microbiol. 11, 517e524. 310e313.
Hull, C.M., Heitman, J., 2002. Genetics of Cryptococcus neoformans. Menkis, A., Jacobson, D.J., Gustafsson, T., Johannesson, H., 2008.
Annu. Rev. Genet. 36, 557e615. The mating-type chromosome in the filamentous ascomycete
Hull, C.M., Raisner, R.M., Johnson, A.D., 2000. Evidence for mating Neurospora tetrasperma represents a model for early evolution
of the “asexual” yeast Candida albicans in a mammalian host. of sex chromosomes. PLoS Genet. 4, e1000030.
Science 289, 307e310. Metin, B., Findley, K., Heitman, J., 2010. The mating type locus
Idnurm, A., Walton, F.J., Floyd, A., Heitman, J., 2008. Identification (MAT ) and sexual reproduction of Cryptococcus heveanensis:
of the sex genes in an early diverged fungus. Nature 451, insights into the evolution of sex and sex-determining chro-
193e196. mosomal regions in fungi. PLoS Genet. 6, e1000961.
Inderbitzin, P., Harkness, J., Turgeon, B.G., Berbee, M.L., 2005. Morran, L.T., Schmidt, O.G., Gelarden, I.A., Parrish 2nd, R.C.,
Lateral transfer of mating system in Stemphylium. Proc. Natl. Lively, C.M., 2011. Running with the Red Queen: host-parasite
Acad. Sci. U. S. A. 102, 11390e11395. coevolution selects for biparental sex. Science 333, 216e218.
King, K.C., Delph, L.F., Jokela, J., Lively, C.M., 2009. The geographic Muller, H., Hennequin, C., Gallaud, J., Dujon, B., Fairhead, C., 2008.
mosaic of sex and the Red Queen. Curr. Biol. 19, 1438e1441. The asexual yeast Candida glabrata maintains distinct a and
King, K.C., Jokela, J., Lively, C.M., 2011. Parasites, sex, and clonal alpha haploid mating types. Eukaryot. Cell 7, 848e858.
diversity in natural snail populations. Evolution 65, 1474e1481. Myosho, T., Otake, H., Masuyama, H., Matsuda, M., Kuroki, Y.,
Kondo, M., Nanda, I., Hornung, U., Schmid, M., Schartl, M., 2004. Fujiyama, A., Naruse, K., Hamaguchi, S., Sakaizumi, M., 2012.
Evolutionary origin of the medaka Y chromosome. Curr. Biol. Tracing the emergence of a novel sex-determining gene in
14, 1664e1669. medaka, Oryzias luzonensis. Genetics 191, 163e170.
Kwon-Chung, K.J., 1976a. Morphogenesis of Filobasidiella neofor- Ni, M., Feretzaki, M., Li, W., Floyd-Averette, A., Mieczkowski, P.,
mans, the sexual state of Cryptococcus neoformans. Mycologia Dietrich, F.S., Heitman, J., 2013. Unisexual and heterosexual
68, 821e833. meiotic reproduction generate aneuploidy and phenotypic
Lang, G.I., Murray, A.W., Botstein, D., 2009. The cost of gene diversity de novo in the yeast Cryptococcus neoformans. PLoS
expression underlies a fitness trade-off in yeast. Proc. Natl. Biol. 11, e1001653.
Acad. Sci. U. S. A. 106, 5755e5760. O’Gorman, C.M., Fuller, H., Dyer, P.S., 2009. Discovery of a sexual
Lang, G.I., Botstein, D., Desai, M.M., 2011. Genetic variation and cycle in the opportunistic fungal pathogen Aspergillus fumiga-
the fate of beneficial mutations in asexual populations. Ge- tus. Nature 457, 471e474.
netics 188, 647e661. Phadke, S.S., Feretzaki, M., Heitman, J., 2013. Unisexual repro-
Lee, N., Bakkeren, G., Wong, K., Sherwood, J.E., Kronstad, J.W., duction enhances fungal competitiveness by promoting
1999. The mating-type and pathogenicity locus of the fungus habitat exploration via hyphal growth and sporulation. Eu-
Ustilago hordei spans a 500-kb region. Proc. Natl. Acad. Sci. U. S. karyot. Cell 12, 1155e1159.
A. 96, 15026e15031. Poxleitner, M.K., Carpenter, M.L., Mancuso, J.J., Wang, C.J.,
Lee, S.C., Corradi, N., Byrnes 3rd, E.J., Torres-Martinez, S., Dawson, S.C., Cande, W.Z., 2008. Evidence for karyogamy and
Dietrich, F.S., Keeling, P.J., Heitman, J., 2008. Microsporidia exchange of genetic material in the binucleate intestinal
evolved from ancestral sexual fungi. Curr. Biol. 18, 1675e1679. parasite Giardia intestinalis. Science 319, 1530e1533.
Levin, T.C., King, N., 2013. Evidence for sex and recombination in Ramirez-Zavaleta, C.Y., Salas-Delgado, G.E., De Las Penas, A.,
the choanoflagellate Salpingoeca rosetta. Curr. Biol. 23, Castano, I., 2010. Subtelomeric silencing of the MTL3 locus of
2176e2180. Candida glabrata requires yKu70, yKu80, and Rif1 proteins.
Liew, W.C., Bartfai, R., Lim, Z., Sreenivasan, R., Siegfried, K.R., Eukaryot. Cell 9, 1602e1611.
Orban, L., 2012. Polygenic sex determination system in ze- Raper, J.R., 1966. Genetics of Sexuality in Higher Fungi. The Ro-
brafish. PLoS One 7, e34397. nald Press Company, New York.
Liew, W.C., Orban, L., 2014. Zebrafish sex: a complicated affair. Roach, K.C., Feretzaki, M., Sun, S., Heitman, J., 2014. Unisexual
Brief Funct. Genomics 13, 172e187. reproduction. Adv. Genet. 85, 255e305.
Lin, X., Hull, C.M., Heitman, J., 2005. Sexual reproduction between Roach, K.C., Heitman, J., 2014. Unisexual reproduction reverses
partners of the same mating type in Cryptococcus neoformans. Muller’s ratchet. Genetics 198, 1059e1069.
Nature 434, 1017e1021. Rodriguez-Carres, M., Findley, K., Sun, S., Dietrich, F.S.,
Lin, X., Huang, J., Mitchell, T., Heitman, J., 2006. Virulence attri- Heitman, J., 2010. Morphological and genomic characteriza-
butes and hyphal growth of C. neoformans are quantitative tion of Filobasidiella depauperata: a homothallic sibling species
traits and the MATa allele enhances filamentation. PLoS of the pathogenic cryptococcus species complex. PLoS One 5,
Genet. 2, e187. e9620.
Lin, X., Litvintseva, A.P., Nielsen, K., Patel, S., Floyd, A., Saul, N., Krockenberger, M., Carter, D., 2008. Evidence of recom-
Mitchell, T.G., Heitman, J., 2007. aADa hybrids of Cryptococcus bination in mixed-mating-type and a-only populations of
Evolution of sexual reproduction 117

Cryptococcus gattii sourced from single eucalyptus tree hol- Wendte, J.M., Miller, M.A., Lambourn, D.M., Magargal, S.L.,
lows. Eukaryot. Cell 7, 727e734. Jessup, D.A., Grigg, M.E., 2010. Self-mating in the definitive
Schaffer, A., 2007. Pas de deux: Why are There Only Two Sexes? host potentiates clonal outbreaks of the apicomplexan para- sites Sarcocystis neurona and Toxoplasma gondii. PLoS Genet. 6,
pas_de_deux.html. e1001261.
Schurko, A.M., Logsdon Jr., J.M., 2008. Using a meiosis detection Whittle, C.A., Votintseva, A., Ridout, K., Filatov, D.A., 2015. Recent
toolkit to investigate ancient asexual “scandals” and the evo- and massive expansion of the mating-type specific region in
lution of sex. BioEssays 30, 579e589. the smut fungus Microbotryum. Genetics 199, 809e816.
Singh, D.P., Saudemont, B., Guglielmi, G., Arnaiz, O., Gout, J.F., Wickes, B.L., Mayorga, M.E., Edman, U., Edman, J.C., 1996.
Prajer, M., Potekhin, A., Przybos, E., Aubusson-Fleury, A., Dimorphism and haploid fruiting in Cryptococcus neoformans:
Bhullar, S., Bouhouche, K., Lhuillier-Akakpo, M., Tanty, V., association with the a-mating type. Proc. Natl. Acad. Sci. U. S.
Blugeon, C., Alberti, A., Labadie, K., Aury, J.M., Sperling, L., A. 93, 7327e7331.
Duharcourt, S., Meyer, E., 2014. Genome-defence small RNAs Wilson, C.A., High, S.K., McCluskey, B.M., Amores, A., Yan, Y.L.,
exapted for epigenetic mating-type inheritance. Nature 509, Titus, T.A., Anderson, J.L., Batzel, P., Carvan 3rd, M.J.,
447e452. Schartl, M., Postlethwait, J.H., 2014. Wild sex in zebrafish: loss
Srikantha, T., Lachke, S.A., Soll, D.R., 2003. Three mating type-like of the natural sex determinant in domesticated strains. Ge-
loci in Candida glabrata. Eukaryot. Cell 2, 328e340. netics 198, 1291e1308.
Statzell-Tallman, A., Belloch, C., Fell, J.W., 2008. Kwoniella man- Wong, S., Fares, M.A., Zimmermann, W., Butler, G., Wolfe, K.H.,
groviensis gen. nov., sp.nov. (Tremellales, Basidiomycota), a tele- 2003. Evidence from comparative genomics for a complete
omorphic yeast from mangrove habitats in the Florida sexual cycle in the ’asexual’ pathogenic yeast Candida glabrata.
Everglades and Bahamas. FEMS Yeast Res. 8, 103e113. Genome Biol. 4, R10.
Sun, S., Billmyre, R.B., Mieczkowski, P.A., Heitman, J., 2014. Uni- Yun, S.H., Arie, T., Kaneko, I., Yoder, O.C., Turgeon, B.G., 2000.
sexual reproduction drives meiotic recombination and Molecular organization of mating type loci in heterothallic,
phenotypic and karyotypic plasticity in Cryptococcus neofor- homothallic, and asexual Gibberella/Fusarium species. Fungal
mans. PLoS Genet. 10, e1004849. Genet. Biol. 31, 7e20.
Tscharke, R.L., Lazera, M., Chang, Y.C., Wickes, B.L., Kwon- Yun, S.H., Berbee, M.L., Yoder, O.C., Turgeon, B.G., 1999. Evolution
Chung, K.J., 2003. Haploid fruiting in Cryptococcus neoformans is of the fungal self-fertile reproductive life style from self-sterile
not mating type a-specific. Fungal Genet. Biol. 39, 230e237. ancestors. Proc. Natl. Acad. Sci. U. S. A. 96, 5592e5597.
Umen, J., Heitman, J., 2013. Evolution of sex: mating rituals of a Zhou, Q., Zhang, J., Bachtrog, D., An, N., Huang, Q., Jarvis, E.D.,
pre-metazoan. Curr. Biol. 23, R1006eR1008. Gilbert, M.T., Zhang, G., 2014. Complex evolutionary trajec-
Vergara, D., Lively, C.M., King, K.C., Jokela, J., 2013. The tories of sex chromosomes across bird taxa. Science 346, 1332.
geographic mosaic of sex and infection in lake populations of
a New Zealand snail at multiple spatial scales. Am. Nat. 182,