S2W3 - Intracellular Compartments and Protein Sorting Flashcards
the volumes taken up by organelles will…
differ for different cell types
percentage volume taken up by cytosol
- half the cell volume
- part site of protein synthesis and degradation
- where many metabolic pathways and cytoskeleton occur
rough ER
- over 50% of total cell membrane
- membrane-bound ribosomes
- synthesis of soluble proteins and transmembrane proteins for the endomembrane
smooth ER
- doesn’t have membrane bound ribosomes
- phospholipid synthesis, detoxification
percentage of total cell membrane of rough ER membrane in liver hepatocyte vs pancreatic exocrine cell
60% in pancreas, 35% in liver
- pancreas has to synthesise enzymes.
why are mitochondria so abundant in liver hepatocytes?
provide energy to support the many metabolic functions on the liver
definition of an organelle
discrete structure or subcompartment of a eukaryotic cell that is specialised to carry out a particular function (most are membrane-enclosed)
how were organelles discovered?
via visualisation in a light or electron microscope
examples of membrane-enclosed organelles
- nucleus
- endoplasmic reticulum
- Golgi apparatus
examples of organelles that are not membrane-bound
- nucleolus
- centrosome
also known as bimolecular condensates
what is protein sorting?
- proteins are nuclear encoded
- mRNA arrives in cytoplasm and translation starts on ribosomes in cytosol
- cytosolic protein doesn’t have a sorting signal, so its default location is the cytosol
- some proteins have a sorting signal called a signal sequence
what does a signal sequence consist of?
a stretch of amino acid sequence in a protein which directs the protein to the correct compartment
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
post-translational protein sorting
- proteins are nuclear-encoded
- fully synthesised in cytosol before sorting
- folded: nucleus, peroxisomes
- unfolded: mitochondria, plastids
co-translational sorting
- proteins are nuclear-encoded
- they have an ER signal sequence and are associated with ER during protein synthesis in the cytosol
proteins that are intended for the nucleus have a
nuclear localisation signal (NLS) for import into the nucleus.
function of the nuclear import receptors
nuclear import receptor (sorting receptor) binds the NLS and move it into the nucleus. nuclear pores act as gates to the nucleus - proteins with the nuclear import receptor are recognised.
function of transcription activators
required in the nucleus for eukaryotic transcription - imported through the nuclear pore and binds to DNA to bind to the activated target gene
function of peroxisomes
contain enzymes for oxidative reactions
- detoxify toxins, break down fatty acid molecules
enzymes imported into the peroxisome through
a transmembrane complex - there is a peroxisomal import receptor (sorting receptor) which binds to the peroxisome import sequence (signal sequence)
how and why do proteins have to move into mitochondria/chloroplasts
- have own genomes and ribosomes
- but most proteins for these organelles are nuclear-encoded
- translated in cytosol and targeted by a signal sequence for import
- proteins are unfolded for import by association with hsp70 chaperone proteins
transmembrane complexes in mitochondria
needs proteins to be unfolded to pass through. signal sequence bound to sorting receptor, which brings it to transmembrane complexes - hsp70 proteins come off as the protein moves through. then the mitochondrial hsp70 binds to it in the mitochondrial matrix (these help the protein fold and remove the signal)
why do proteins sort to the ER?
entry point to endomembrane system (ER, Golgi, endossâmes, lysosomes, or up to plasma membrane)
what is co-translational translocation?
insertion of protein into ER starts as translation continues (whole ribosomal complex moves together)
ER signal sequence is usually at
N-terminus
types of proteins entering the ER:
- soluble proteins
- transmembrane proteins (eg channel)
are ribosomes specific to cytosol/ER?
no; once they are done translating protein, they move back to the cytosol and pick up any protein (no specificity)
is ER signal hydrophobic or hydrophilic?
hydrophobic
describe the process for co-translational translocation of a soluble protein
- translation starts, N-terminal ER signal sequence emerges
- recognised by SRP, elongation arrest by SRP
- SRP-ribosome complex binds to SRP receptor and moves it to the translocon
- translocon opens
- protein synthesis resumes with protein transfer into ER lumen
- signal peptidase cleaves ER signal sequence, which is hydrophobic - in lipid bilayer
- protein released into ER lumen
- translocon closes
destination of soluble protein
lumen of an endomembrane organelle or secretion at PM
translocon
protein channel complex in the endoplasmic reticulum (ER) membrane that facilitates the translocation of proteins across or into the ER membrane
co-translational translocation for transmembrane protein (N-terminal ER signal sequence)
- translation starts, N-terminal ER signal sequence emerges
- recognised by SRP, elongation arrest by SRP
- SRP-ribosome complex binds to SRP receptor which moves it to the translocon
- translocon opens
- protein synthesis resumes with protein transfer into ER lumen
- stop-transfer sequence, which has an internal hydrophobic segment (single-pass transmembrane protein w a membrane spanning alpha helix) enters translocon
- protein transfer stops and transmembrane domain released into lipid bilayer
- signal peptidase cleaves ER signal sequence and translocon closes
- N-terminus is in the ER lumen, C-terminus is in the cytosol, hydrophobic sequence is in it
co-translational translocation for transmembrane protein (internal ER signal sequence)
- translation starts, internal start-transfer sequence emerges
- recognised by SRP, elongation arrest by SRP
- SRP-ribosome complex binds to SRP receptor which moves it to the translocon
- translocon opens
- protein synthesis resumes with protein transfer into ER lumen
- stop-transfer sequence enters translocon
- protein transfer stops
- start-transfer sequence and stop-transfer sequence (internal hydrophobic segments = membrane spanning alpha helices) are released into lipid bilayer
- translocon closes
destination of transmembrane protein
membrane of an endomembrane organelle or in the plasma membrane
N-terminal ER signal sequence
- stretch of hydrophobic amino acids at N-terminus of protein
- removed by signal peptidase
Internal ER signal sequence
- stretch of hydrophobic amino acids (start-transfer sequence)
- not removed - remains part of protein = membrane spanning alpha helix
- In double-pass or multi-pass transmembrane proteins, these internal sequences work with stop-transfer sequences to embed multiple segments of polypeptides within membranes.
similarity and differences between N-terminal and internal ER signal sequence
both types of sequences direct proteins to enter or integrate into the ER membrane via similar mechanisms involving SRP recognition and targeting to translocons, their positions within polypeptides and subsequent fates differ.
differences between hydrophobic stop transfer and start transfer sequences
no differences other than order in the protein
what components do intracellular compartments in the endomembrane system exchange?
lipids and proteins
secretory pathway
proteins and lipids made in the ER are delivered to other compartments
- ER to outside (exocytosis)
- ER to lysosomes (via endosomes)
endocytic pathway
contents moved into the cell (endocytosis)
retrieval pathway
retrieval of lipids/selected proteins for reuse
what happens to the individual leaflets of the membranes during endo/exocytosis?
maintain their orientation throughout
vesicular transport
- vesicle: small, membrane-enclosed organelle in cytoplasm of a eukaryotic cell
- shuttle components back an forth in the endomembrane system (eg ER to Golgi)
- two main components: soluble proteins inside the vesicle and transmembrane proteins embedded in membrane of vesicle
cargo proteins
receptors can be used to select for cargo proteins and then released into the ER lumen
constitutive exocytosis pathway
- in all eukaryotic cells
- continual delivery of proteins (transmembrane, soluble) and lipids to plasma membrane
- includes constitutive secretion of soluble proteins (eg collagen for ECM)
regulated exocytosis pathway
- regulated secretion - in specialised cells
- stored in specialised secretory vesicles
- extracellular signal leas to vesicle fusion with plasma membrane and contents are released
- eg pancreatic β cells (insulin release with increased blood glucose)
path of a secreted protein from translation to plasma membrane
Translation starts on cytosolic ribosomes
* ER Signal sequence at N-terminus directs protein to ER
Co-translational translocation at ER
* Protein inserted through ER membrane by a translocon protein
* ER Signal sequence cleaved and left behind in ER membrane
* Secreted protein ends up in ER lumen
Secreted protein
* Moves in transport vesicles through secretory pathway (ER → Golgi apparatus → Plasma membrane)
* Vesicle membrane fuses with plasma membrane during exocytosis
* Secreted protein released to extracellular spac
Path of a transmembrane protein from translation to plasma membrane
Translation starts on cytosolic ribosomes
* ER Signal sequence (N-terminal or Internal) directs protein to ER
Co-translational translocation at ER
* Protein inserted through ER membrane by a translocon protein
* There are different ways that a transmembrane protein can be
inserted into ER membrane
Transmembrane protein
* Moves in transport vesicles through secretory pathway (ER → Golgi apparatus → Plasma membrane)
* Vesicle membrane fuses with plasma membrane during exocytosis
* Transmembrane protein transferred to plasma membrane
how is maintenance of membrane protein asymmetry maintained?
- each membrane protein has a specific orientation
- this is a result of membrane orientation in the ER
- this protein symmetry is maintained through vesicular transport
Golgi function
receives proteins and lipids from the ER, modifies them, and then dispatches them to other destinations in the cell
Golgi structure
stack of flattened membrane-enclosed stacks (cisternae)
enters via:
- cis Golgi network
- cis cisternae
- medial cisterna
- trans cisterna
- trans Golgi network
leaves
animal vs plant cell Golgi
animal cells tend to have one bind Golgi structure; plant cells have a lot of smaller Golgi spread out throughout the cell
protein glycosylation
- starts in the ER
- a single type of oligosaccharide is attached to many proteins
- Complex oligosaccharide processing occurs in the Golgi apparatus (a multistage processing unit - different enzymes in each cisterna)
- glycosylation modifications for proteins and lipids (glycosylated lipids and proteins end up on the outside of cell to protect membrane + proteins from damage)
endocytic pathway: endosomes and lysosomes
digest material that is no longer needed by the cell
Endosomes
- membrane-bound organelles
- contain material ingested by endocytosis
- endocytic vesicles fuse to early endosomes and ingested material sorted so that it will either go to the Golgi (recycling pathway) or to lysosomes (degradation pathway)
Degradation:
2. early endosomes mature into late endosomes
3. lysosomal proteins (hydrolases, H+ pump) continue to be delivered from trans Golgi network to either late or early endosomes
4. late endosomes mature into lysosomes
lysosomes
- membrane-bound organelles
- contain hydrolytic enzymes to digest worn-out proteins, organelles, other wast
difference between early and late endoscope
late not recycling cargo anymore, committed to cargo degradation pathway
where does digestion start?
at the late endosome
how does trans Golgi know that it needs to package up lysosomal proteins into vesicles to send them to vesicles?
specific types of sugar groups can be added to vesicles that are signals
types of enzymes contained in lysosomes
around 40 types of hydrolytic enzymes known as acid hydrolases (proteases, nucleases, lipases, etc)
lysosomes are acidified by
- H+ pump (V-type ATPase)
- low pH needed for hydrolytic enzymes
how does the lysosome protect the rest of the cell from digestion?
- it is membrane bound
- lysosomal membrane proteins (noncytosolic face, inside) are glycosylated for protection from proteases
function of transport proteins in lysosomal membrane
transfer digested products to cytosol (amino acids, sugars, nucleotides)
why do cytosolic proteins not have signal sequences?
as they stay in the cytosol`
3 different pathways of protein movement
secretory pathway
endocytic pathway
retrieval pathway
what gives vesicles directionality?
- directed movement of transport vesicles
- pulled by motor proteins associated with cytoskeleton
