Intracellular Compartments

and Protein Sorting

Reading: Alberts et al. 5th edition
695-704, 721-727, 766-769, 779-783

Intracellular Compartments

Cytosol

- half the cell volume

- protein synthesis and degradation
- intermediary metabolism (degradation and synthesis of small molecules)

Table 12-1 Molecular Biology of the Cell ( Garland Science 2008)

Definition: organelle
A subcellular compartment or large macromolecular
complex, often membrane-enclosed, that has a
distinct structure, composition, and function.
Examples of membrane-enclosed organelles:
- Nucleus
- Endoplasmic Reticulum
- Golgi Apparatus
Examples organelles that are not membrane-bound:
- Nucleolus
- Centrosomes
3

(endocytosis)
(Digestive Enzymes)
(modifies proteins & lipids)

(oxidative
reactions)

(protein synthesis)

(ATP synthesis)
(ER)
(RNA, DNA synthesis)

RER synthesis of transmembrane, organellar and secreted proteins

SER phospholipid synthesis, detoxification
Figure 12-1 Molecular Biology of the Cell ( Garland Science 2008)

Figure 12-2 Molecular Biology of the Cell ( Garland Science 2008)

detoxifies

secrete protein

ER usually
50%

Table 12-2 Molecular Biology of the Cell ( Garland Science 2008)

Intracellular Compartments
are Dynamic
They exchange components:
Lipids
Proteins

Figure 12-5 Molecular Biology of the Cell ( Garland Science 2008)

The Endomembrane system

/vacuole (in plants)

Figure 12-1 Molecular Biology of the Cell ( Garland Science 2008)

Organelles of the Endomembrane System:

Endoplasmic Reticulum (ER)
Golgi Apparatus
Endosomes
Lysosomes (Plant Vacuoles)

these components function together in:

the biosynthetic and endocytic pathways
Mitochondria and Chloroplasts are not considered to be part of this system as
they have their own specific functions.
What about Peroxisomes?
- Controversial, have a dual origin
- Components derived from both the ER and cytosol (figure 12-33)
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A) Biosynthetic/Secretory
pathways
Biosynthesis
Proteins and lipids made
in the ER delivered to the
membranes of other
organelles

Secretion
Secreted proteins
Contents move out of the
cell (Exocytosis)

B) Endocytic Pathway
Contents move into the
cell (Endocytosis)
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A) exocytosis:

vesicle contents
delivered to extracellular space
vesicle membrane becomes part of the PM

B) Endocytosis:

PM (plasma membrane) forms the vesicle membrane

vesical luminal
contents come from extracellular space

Figure 13-1 Molecular Biology of the Cell ( Garland Science 2008)

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Vesicular Transport
Vesicle: small, membrane-enclosed
organelle in the cytoplasm of a
eukaryotic cell
They shuttle components back and
forth in the endomembrane system
For example:

from the ER to the Golgi

goes from interior to interior

Luminal leaflet
Cytosolic leaflet
Figure 12-7 Molecular Biology of the Cell ( Garland Science 2008)

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Lets follow the path of a secreted protein

from translation to secretion. Ribosome
mRNA

mRNA arrives in the

cytoplasm and translation
starts on ribosomes in the
cytosol.
While translation is still
occurring insertion of the
protein into the ER starts

Figure 12-35a Molecular Biology of the Cell ( Garland Science 2008)

Signal sequence

cytosol
ER Lumen

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hydrophobic

Figure 12-38 Molecular Biology of the Cell ( Garland Science 2008)

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Pancreatic cells make lots of secreted

proteins
Use these cells to follow the
path of newly synthesized
proteins.
Provide the cells with a short
pulse of radioactive amino
acids (AA).
Follow the path of these AA
as they are incorporated into
proteins.
A Technique called a:
pulse-chase experiment

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Proteins move from the...

RER

(chase time)

Golgi
Apparatus

(chase time)

Secretory
Vesicles

(chase time)

red dots are labeled proteins

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Secretory Pathway
Constitutive Pathway:
Continual production of
secreted proteins.
e.g. collagen

Regulated Pathway:
Proteins are stored in
secretory granules ready
for export in response to
a stimulus.
e.g. Neurotransmitter
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Protein Sorting Mechanisms

1. Gated: proteins move between
the cytosol and nucleus
through NPCs
- nuclear pore complexes
1. Transmembrane:
protein translocators needed to
transport specific proteins
across a membrane.
3. Vesicular: membrane-enclosed
transport vesicles ferry proteins
from one compartment to
another.
Figure 12-6 Molecular Biology of the Cell ( Garland Science 2008)

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Signal Sequences
A stretch of the amino acid sequence of a protein
that directs the protein to the correct location in
the cell.
Each signal sequence specifies a specific
destination in the cell.
Specific signal sequences direct proteins to nucleus,
mitochondria, ER, peroxisomes, etc.

Signal sequences are recognised by sorting

receptors that take proteins to their destination.
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Signal Sequences
Often found at the Nterminus of the protein.
Signal peptidases: can
remove the signal sequence
from the finished protein.

NH2

Can be internal stretches of

AA which remain part of the
protein.
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Recap: Sorting of a Secreted Protein.

Translation starts on cytosolic ribosomes.
Signal sequence at the amino-terminal end directs the
protein to the ER.
The signal sequence is: hydrophobic
This is:
Co-translational translocation to ER
Protein inserted through the membrane by a:
protein translocator.
Signal sequence cleaved, left behind in the ER
membrane, secreted protein ends up in the ER lumen.
Secreted protein moves in transport vesicles via the
secretory pathway.
Released by Exocytosis at the Plasma membrane.
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What about a transmembrane protein?

CytosolER

hydrophilic

hydrophobic

Figure 12-46 Molecular Biology of the Cell ( Garland Science 2008)

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Two different ways a single pass transmembrane

protein can be oriented in the ER membrane
(for a protein with an
internal signal sequence)
determined by
amino acids right
by signal sequence
- side on ER lumen
+ side in cytosol

Figure 12-47 Molecular Biology of the Cell ( Garland Science 2008)

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A Multipass Transmembrane Protein

Figure 12-48 Molecular Biology of the Cell ( Garland Science 2008)

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Rhodopsin
(found in rod photoreceptor cells)

Figure 12-49 Molecular Biology of the Cell ( Garland Science 2008)

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Maintenance of
Membrane asymmetry
Each protein is inserted
into the membrane in
the ER in a specific
manner
This protein asymmetry
is maintained through
vesicular transport
a result of the process of
membrane insertion
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Lets follow the path of a transmembrane protein

from translation to the plasma membrane.
CytosolER
ER Golgi
Whats going on
in the Golgi?
Glycosylation
lipids and proteins

27

Definitions
The Golgi apparatus receives proteins and
lipids from the endoplasmic reticulum,
modifies them, and then dispatches them to
other destinations in the cell.

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The Golgi Apparatus

Nuclear envelope

ER
usually 4-6 stacks

Figure 13-25a Molecular Biology of the Cell ( Garland Science 2008)

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Protein Glycosylation
Actually starts in the ER: A single type of oligosaccharide is
attached to many proteins.

Complex oligosaccharide processing required for

mature proteins occurs in the Golgi
non-cytosolic side usually glycosilated

Golgi:
a multistage
processing unit
different enzymes
in each cisterna

Figure 10-28b Molecular Biology of the Cell ( Garland Science 2008)

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The Golgi Apparatus is also involved in

Glycosylation of Lipids
Plant cell wall polysaccharides
Pectin and hemicellulose
Extracellular matrix of animal cells
Glycosaminoglycans
Some proteins are sorted back to the ER from the
Golgi
different signal
retrieval pathway
31

Recap: Sorting of a PM protein.

Translation starts on cytosolic ribosomes.
Signal sequence directs the protein to the ER.
There are different ways the protein can be inserted
into the membrane.
The signal sequence is: hydrophobic
This is:
Co-translational translocation to ER
Protein inserted through the membrane by a:
protein translocator.
Protein moves in transport vesicles to the PM
Vesicle membrane fuses with the PM during
Exocytosis, transferring the protein to the PM
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Lysosomes and Endosomes

Figure 13-3b Molecular Biology of the Cell ( Garland Science 2008)

33

Definitions
Lysosome: a small sac of digestive enzymes
that functions in degrading worn-out
organelles, as well as macromolecules and
particles taken into the cell by endocytosis.
Endosome: membrane-bound organelle that
carries materials newly ingested by
endocytosis and passes them to lysosomes for
degradation.
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Phagocytosis/Endocytosis

An amoeba ingesting a food particle

food vacuole=endosome

A leukocyte ingesting a yeast cell

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Lysosomes are the main site of

intracellular digestion
Contain approx 40 types
of hydrolytic enzymes:
Proteases, nucleases,
lipases etc.

These enzymes are:

Acid hydrolases
Require an acidic
environment

lysosomes are acidified by a proton pump

(V-type ATPase)

Figure 13-36 Molecular Biology of the Cell ( Garland Science 2008)

36

Lysosomes
Low pH requirement of lysosomal
enzymes protects contents of the
cytosol from digestion.
Lysosomal membrane proteins are
glycosylated to protect them from
proteases in the lumen
Transport proteins in the lysosomal
membrane transfer the products of
digestion out of the lysosome to the
cytosol
AA, sugars, nucleotides
Figure 13-37 Molecular Biology of the Cell ( Garland Science 2008)

37

Formation of lysosomes
Lysosomal proteins from the ER and Golgi
are incorporated into endosomes at different stages.
classical lysosomes:
Majority of endocytosed
material has been digested

However lysosomal
hydrolases delivered
from the Golgi are found in
early and late endosomes
Where do the membrane lipids of lysosomes come from?
Figure 13-3b Molecular Biology of the Cell ( Garland Science 2008)

38

Lysosome proteins are found in

heterogeneous looking compartments

lysosomal proteins are delivered to endosomes directly from the Golgi in

vesicles
Mature lysosomes also fuse with endosomes to recycle lysosomal
proteins

Figure 13-38 Molecular Biology of the Cell ( Garland Science 2008)

39

Plant Vacuoles
Occupy 30-90% of the cell volume
Diverse
Involved in:
Digestion
Nutrient storage
Waste storage
Increasing cell size
Turgor pressure
Remember how osmosis affects plant cells!
Figure 13-39 Molecular Biology of the Cell ( Garland Science 2008)

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A large increase in
volume of a plant cell
without increasing
the volume of the
cytosol

Figure 13-40 Molecular Biology of the Cell ( Garland Science 2008)

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Peroxisomes

catalase uses H2O2 to oxidize compounds, peroxidation reaction

Use molecular oxygen to

oxidise organic molecules
Oxidative reactions
produce hydrogen
peroxide (H2O2)
H2O2 is toxic; also
degraded in peroxisomes
Function in the
breakdown of long fatty
acid chains.
Figure 12-30 Molecular Biology of the Cell ( Garland Science 2008)

42

Peroxisomes contribute to the

myelination of nerve axons
Schwann cell wraps around the
axon.
Tight layer of Schwann cell PM
wrapped around the axon.
Special composition of PM
phospholipids in Schwann cells.

Plasmalogens most abundant.

Made in peroxisomes
Peroxisomal disorders cause
neurological diseases.
Figure 11-32a Molecular Biology of the Cell ( Garland Science 2008)

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Model of Peroxisome Formation

- Proteins inserted into the ER

- Proteins imported from the cytosol
Figure 12-33 Molecular Biology of the Cell ( Garland Science 2008)

Found in
peroxisomes
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Overview: Protein Sorting (1)

movement of proteins

within cell between

different compartments
out of cell
into cell

protein synthesis initiated on

ribosomes in cytosol
proteins must be sorted to the correct
location signal sequences
Cytosolic proteins stay in the cytosol
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Overview: Protein Sorting (2)

1) Post-translational process
proteins fully synthesized in
cytosol before sorting
Unfolded: mitochondria, plastids
Folded: nucleus, peroxisomes
2) Co-translational process
Endoplasmic Reticulum (ER)
proteins with ER Signal Sequence
associated with ER during
protein synthesis

Nucleus

mRNA
ribosome

protein
synthesis

ER
membrane
46

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How does a nuclear protein get into

the nucleus?
Transport is:
Gated
Occurs through:
Nuclear pore complexes
(NPC)
This process can be regulated
by intracellular signals.
e.g. Ca2+, hormones
Figure 12-8 Molecular Biology of the Cell ( Garland Science 2008)

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Protein Sorting: Gated Transport

Proteins moving between cytosol and nucleus
Nuclear Pore Complex
selective transport
of macromolecules
free diffusion of

small molecules
(<5,000 daltons)

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Figure 12-10 Molecular Biology of the Cell ( Garland Science 2008)

Proteins Moving between the Cytosol and Nucleus

Transport of cargo through Nuclear Pore
Complex (NPC)
transport in both directions
1) Nuclear Import
from cytosol to nucleus
2) Nuclear Export
from nucleus to cytosol

Page 704 Molecular Biology of the Cell 5/e

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Nuclear Import Signal Sequences

a protein with an intact nuclear

transport singal is found in nucleus

Figure 12-11 Molecular Biology of the Cell ( Garland Science 2008)

when signal mutated protein stays in

cytosol

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Remember....

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Estrogen Receptor is a ligandmodulated regulator of transcription

When estradiol is NOT present
estrogen receptor -> cytosol
When estradiol enters the cell
binds estrogen receptor
Ligand-bound ER moves into nucleus
through NPCs
Binds to enhancer sites in the genome
and activates transcription of
target genes

52

Protein Sorting: Mechanisms

gated transport

transmembrane
transport
requires protein
translocators

vesicular transport
vesicles move protein
between compartments

53

What gives the vesicles directionality?

Directed movement of transport vesicles,
pulled by motor proteins associated with the
cytoskeleton

54

End

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