vesicular trafficking from ER -> Golgi -> ER Flashcards
ER exit site (ERES)
ER subdomain involved in the formation of transport vesicles destined for the Golgi
- usually located next to the cis Golgi
- proper packaging of vesicles with the correct cargo destined for the golgi
COP proteins
- a layer of soluble coat proteins attached to the outer surface of the transport vesicles
2 main functions…
1. recognize and concentrate specific components to be incorporated into the budding vesicle
2. mediate ERES membrane curvature and formation of the budding vesicle
3 major classes of coat proteins
COP II: moves anterograde direction, from ERES to Golgi
COP I: moves retrograde direction, from Golgi to ERES OR backwards within the golgi
Clathrin: moves anterograde and retrograde, from Golgi OR PM to endosomes
what is Sar1 protein
- A G-protein recruited to the cytosolic surface of the ERES responsible for COP II vesicle formation
how does Sar1 mediate COP II transport vesicle formation
- Sar1-GDP is recruited to the ERES and binds to Sec 12 (a GEF) which generates Sar1-GTP (active)
- Sar1-GTP gets integrated into the outer leaflet of the ERES which causes the ERES membrane to curve outwards
- Sar1 recruits Sec23 and 24 to the ERES membrane to promote further curvature, followed by Sec13 and 31 to act as the outer scaffold coat for the vesicle
Vesicle targeting and fusion at the CGN
- recognition of the incoming vesicle at the recipient membrane is mediated by Rab proteins. Rabs associate with the membrane in a GTP dependent manner
- docking of the vesicle at target membranes is mediated by SNARE proteins. specific v-SNAREs interact with t-SNAREs to form a SNARE complex
- fusion of the vesicle and target membranes, allows for release of vesicle proteins into lumen of acceptor
- disassembly of SNARE complexes
what are Rab proteins
- responsible for recognition of incoming vesicles and recipient membranes in vesicle transport to CGN
- RAB-GTP gets incorporated into vesicles and when the vesicle fuses at CGN membrane RAB-GTP is incorporated into it
what are SNARE proteins
- responsible for docking of vesicles at target membranes (especially at CGN)
- cytosolic facing domains are involved in SNARE-SNARE protein binding
- together Rabs and SNAREs contribute to specificity of vesicle targeting
- SNARE complex pulls vesicle and recipient membranes closer together
2 classes of SNARE proteins
v-SNAREs: located on transport vesicles
t-SNAREs: located on acceptor membranes
What is the fate of vesicle-specific proteins or proteins that “escape from the ER?
- returned to the ER from the CGN by specific retrieval signals
- transported back in COP II vesicles
what is the purpose of retrograde transport from CGN to ER
- serves to recycle membrane and return v-SNAREs and Rab proteins that serve in COP II vesicle transport
- uses v and t-SNAREs, as well as Rab and Rab effectors to prevent fusion of COP II vesicles with the EER membrane
process of ER protein retrieval involving a KDEL sequence
- resident soluble ER proteins possess a C-terminal KDEL sequence (retrieval sequence)
- escaped ER proteins in the CGN are recognized by the KDEL receptor (in the CGN)
- the luminal facing domain of the KDEL receptor binds to the KDEL sequence of the escaped protein
- the cytosolic-facing domain of the KDEL receptor binds to COP I components
- COP I coated vesicles return the ER protein-KDEL receptor complex back to the ER
- at the ER, KDEL receptor releases the resident ER protein and returns to the CGN
what role does pH play when returning ER resident proteins from the CGN
in the CGN, higher [H+] which allows protein with KDEL sequence to bind the KDEL receptor
in the RER, lower [H+] which allows the KDEL receptor to release the ER-resident protein
retrograde transport and KKxx
used for the transport of escaped ER membrane proteins
- most resident ER membrane proteins have a cytoplasmic facing C-terminal dilysine (KKxx) sequence
- KKxx serves as an ER retrieval sorting signal
- proteins that cycle between the ER and CGN contain the KKxx retrieval signal
- KKxx is recognized by COP I at the CGN, COP I coated vesicles bud off the CGN and dock/fuse with the ER
structure of the golgi complex
- contains membrane-bound cisternae
- CGN - cis cisternae - medial cisternae - trans cisternae - TGN
Golgi matrix
mediates the organization of the Golgi complex (stack)
- consists of golgi peripheral and integral membrane proteins
- gold matrix proteins link Golgi to the cytoskeleton
GRASPs
- serve as tethering proteins to link different golgi sub-compartments together
- depletion of GRASP complex results in disassembly of the golgi complex
GMAP-210
facilitates ER to cis-Golgi communication via a direct interaction with the microtubules between these 2 organelles
Golgins
- long filamentous proteins that tether parts if the golgi to the cytoskeleton and other sub cellular compartments
- dimerize and are regulated by G-proteins to anchor them to the Golgi membrane
- if not expressed, the golgi would lose communication with the cytoskeleton and other sub cellular compartments for transport and communication
what can cause the golgi to become disordered
- lack of solid interaction with the cytoskeleton (lack of golgins and GRASPs)
Cis golgi network
- the destination for material from the ER
- has interconnected tubules and vesicles adjacent to the ERES
- site of COP I vesicle assembly for retrograde transport to the ER
- site of anterograde transport from CGN to the rest of the golgi
- destination of COP I vesicles from cis cisternae
Golgi cisternae
- 3 or more large, flattened cisternae divided into cis, medial and trans
- location of golgi metabolism
Trans golgi network
- serves as a sorting station: involved in anterograde transport of materials from golgi to other compartments
- site of clathrin vesicle assembly for transport to endosome or lysosome
- site of secretory vesicle assembly for transport to the plasma membrane
- site of COP I vesicle assembly for transport back to trans cisternae
functions of the golgi complex
- processing plant of the cell
- synthesis of complex polysaccharides
- modification of proteins and lipids (e.g. glycosylation)
- transport and sorting of proteins
- sorting of proteins typically occurs at CGN or TGN
Glycosylation at the Golgi
- N-linked and O-linked
- cis, medial and trans golgi cisternae possess unique glycosyltransferase enzymes which sequentially modify the oligosaccharide core
N-linked: synthesis begins in ER, modified in golgi
O-linked: synthesis/modification entirely in Golgi
Modifications to target proteins in Golgi to the lysosome
Step 1
- in the cis Golgi cisternae, mannose on the core oligosaccharide is phosphorylated.
- GlcNAc phosphotransferase recognizes a signal patch in the protein meant to be targeted to the lysosome
- N-acetyl glucosamine-1-phosphate is transferred to a specific mannose residue
Step 2
- in the medial Golgi, N-acetlyglucosamine group is removed by phosphodiesterase
- the lysosomal enzyme now contains mannose-6-phosphate (identifying feature!)
- M6P is recognized by M6P receptors in the TGN and the enzyme is sorted to lysosomes
pH-dependent delivery of lysosomal proteins
- M6P receptor/lysosomal enzyme is incorporated into clathrin-coated lysosomal vesicles and targeted to lysosome
- pH > 6 in the Golgi which allows lysosomal protein to bind M6P receptor
- pH < 6 in the lysosome which causes the protein to be released from the M6P receptor
