ER/Golgi transport Flashcards
Golgi order
- Cis-network
- Cis
- Medial
- Trans
- Trans-network (TGN)
Possible destination from the Trans-Golgi-Network (TGN)
—> late endosome —> lysosome
—> constitutive secretion
—> regulated secretion
(Retrograde) to trans-Golgi and beyond
As the golgi cisternae move to their new location…the mature through ?
Retrograde movement of enzymes needed for an earlier compartment
Anterograde movement =
Retrograde movement =
- Cisternal maturation
2. Retrograde vesicles
Three different types of vesicles
- COPII (anterograde)
- COPI (retrograde)
- Clathrin/adapter protein (TGN —> endosome/lysosome & plasma membrane —> endosome)
GTPase for COPII vesicles
Sar1
GTPase for COPI vesicles
ARF
GTPase for all non-COPII vesicles
ARF
= COPI + all clathrin vesicles
Trans-golgi —> endosome
Adapter protein?
Clathrin + AP1 complexes (or GGA)
Plasma membrane —> endosome
Adapter proteins?
Clathrin + AP2 complexes
Golgi —> lysosome, melanosome, or platelet vesicles
Adapter proteins?
AP3 complexes
How are integral proteins carried in COPII vesicles from ER —> cis-Golgi
Soluble proteins?
Integral = inserted in the vesicle membrane
Soluble = bind to specific ER membrane-bound receptors
What does the COPII coat protein bind to?
The cytosolic domains of the integral membrane cargo
The receptor for the soluble cargo
The cytosolic domains of the ER membrane…curved nature gives the vesicle a round shape
What happens once the formation of COPII vesicle is complete and pinches off the ER?
The vesicle starts to de-coat (lose the COPII proteins)
Exposes the v-SNAREs proteins in the vesicle membrane
Role of the v-SNAREs
ER —> cis-golgi
Bind to t-SNAREs (target SNAREs)
Mediating vesicle fusion
At this point…
The integral cargo is now integral to the cis-membrane
The soluble cargo is bound to its receptor, bound to the cis-membrane
COPI transport
Same mechanism as COPII
Just in reverse direction and different coat protein
Still uses v and t-SNAREs
GTPase function in vesicle transport
- binds GTP and hydrolyzes the GTP —> GDP
COPI = ARF
COPII = Sar1
Two important functions
- Vesicle formation
- De-coating
GTP-bound form of GTPase (Sar1 or ARF)
N-tail of protein extends into the vesicle membrane (hydrophobic interactions)
Can interact with coat proteins (COPI/II)
Therefore…Sar1 or ARF aids in
- The binding of the coat proteins
- Recruitment of cargo (indirectly)
- Inherent curvature of vesicle
GTP hydrolysis by GTPase
Promotes conformational change of GTPase
Retracts N-tail out of membrane
Lessens the coat proteins association with the vesicle membrane
Mediates coat dissassmebly
Processing of N-linked oligosaccharide complex
Cis golgi vs. trans golgi
Cis = high mannose glycosylation
Trans = complex glycosylation
**proteins can be hybrids of both
High mannose modifications can be enable proteins to be targeted by ?
Lysosomes
High mannose gylcosylation
(Man)8(GlcNAc)2 —> (Man)5(GlcNAc)2
**lose 3 mannose
In TGN,
To get proper cargo into the correct type of transport vesicle…
The adapter protein (AP) binds a cytosolic domain sorting signal on the cargo (or cargo receptors)
The AP also interactes with the coat protein (usually Clathrin)
What transport from the TGN would not have clathrin as the coat protein?
Retrograde from the TGN —> trans-golgi
COPI
Cargo signal for TGN —> lysosome transport
Mannose-6-phosphate (M6P)
I-Cell Disease
General/symptoms
General theme: mis-targeting of proteins from the TGN
Symptoms: 1yr old coarse facial features, craniofacial abnormalities, psychomotor retardadtion, stunted growth
M6P modification sorting signal
TGN —> lysosome transport
Occurs on one of the mannose sugars on the N-linked oligosaccharyl high mannose complex
TGN —> lysosome transport
Steps
- M6P lysosomal proteins packaged into AP3 containing vesicles (M6P receptor)
- De-coats
2A. Clathrin and AP3 are recycled back
- Uncoated vesicle fuses with late endosome/lysosome
- Low pH in lysosome, acid hydrolase (M6P protein) now active
4A. M6P receptor is recycled back to TGN
- Or to the plasma membrane…in case a hydrolase is accidently released out of cell
Mechanism of I-cell Disease
Lysosomal proteins are not tagged with M6P
Therefore…all of lysosomal proteins are do not go to lysosomes…but instead are constitutively secreted out of the cell
Now lysosomes cannot break down substrates
How the M6P cargo signal is created in the cis-golgi
UDP-GlcNAc (substrate) used to add GluNAc-phosphate to one of the mannose sugars
Enzyme #1 = GlcNAc phosphotransferase
Then GlcNAc is removed to leave the M6P sorting signal
Enzyme #2 = phosphodiesterase
What enzyme is blocked by I-Cell Disease
GlcNAc phosphotransferase
Prevents any M6P from being made
Causes a build up of multiple types of inclusion bodies within the lysosome (mainly carbs and lipids)
what is I-cell disease referred to as ?
Mucolipidosis (ML-II)
Characteristics of mucopolysaccharidoses and sphingolipidoses (both types of LSDs)
ML-I
Sialidase deficiency
ML-III
Pseudo-Hurler Polydystrophy
Less severe case of I-cell disease (same enzyme)
What is the root cause of most LSD? And why are ML-II/III different
Single defective hydrolase…so get a buildup of a single class of molecule
ML-II/III have a problem in the targeting part of a lysosomal formation…so many types of molecules build up
MPS-I
Name = Hurler-Scheie syndrome
Enzyme = alpha-L iduronidase
MPS-II
Hunter syndrome
Iduronate-2-sulphatase
MPS-III
Sanfilippo syndrome
Heparan-N-sulphatase
Alpha-N-acetlyglucoaminidase
AcetlyCoA:N-acetyltransferase
N-acetylglucosamine 6 sulphatase
Gaucher disease
Beta-glucosidase
Gangliosidosis type I
Tay Sachs disease
Beta-hexosaminidase A
Gangliosidosis type II
Sandhoff disease
Beta-hexoaminidase A and B
Lysosome pH range
5.0-6.0
Cytosolic pH
7.2-7.4
Endocytosis
Plasma membrane invaginates to form small endocytic vesicles (endosomes)
