Lecture 13 Flashcards
describe vesicle traffic
Multiple donor and acceptor (target) membranes in secretory pathway
Two mechanisms ensure that vesicles transport their contents to the correct acceptor membrane
name the 2 mechanisms for vesicle traffic
Rab GTPase proteins provide specificity of vesicle targeting and attachment to acceptor membrane (Traffic 2)
SNARE fusion proteins provide specificity during fusion of vesicles with acceptor membrane (Traffic 3)
why multiple steps in secretory pathway to the golgi apparatus
if mistake= cannot repair, can go one step back
many ptms providing functions
describe transport through golgi
golgi organized into stack of membranes = cis (closer to er), medial (middle), trans (closer to pm)
3 models
cytosolic protein matrix maintains organization of stack
describe transport through golgi - old model
Vesicles transport cargo between layers
cis–>trans–>membrane binds cop2
describe transport through golgi - new model
each layer matures and becomes next layer
no cop2 vesicles
each compartment matures to next
golgi resident proteins are carried backwards by COP1 vesicles
clathrin coated vesicles carry cargo to pm and endosomes
describe glycosylation in golgi
N-linked glycans are modified by removal of mannoses and addition of different sugars, often with negative charge
Other complex oligosaccharides are attached to Ser and Thr side chains – O-linked glycosylation
Many final combinations of oligosaccharides – heterogeneity
describe glycosylation in golgi generally
diff from er
gives specificity
can modify mannoses, polyglycans, many diff combos, diff types of glycans added
sugar - makes protein more soluble so no aggregation, acts as coat - prevents from degradation
rich in OH so as progress = diff combos of signalling pathways
helps proteins fold properly
what are the functions of glycosylation in the golgi
promotes protein folding
Sugars have limited flexibility ->Protects from proteases, stabilizes protein structure (protective coat)
Signaling hubs (regulation of development
describe how glycosylation in golgi promotes protein folding
Makes folding intermediates more soluble (prevents aggregation)
Sequential modifications -> Glyco-code ->Progression folding or degradatio
describe modifications in the golgi - proprotein convertases
som pm and extracellular proteins are made as longer inactive form at the er, then cut by proprotein convertases into a shorter, active form at the golgi
proteases recognize specific patterns of aas - endonucleolytic cleavage and liberates active part of protein
cleavage often activates proteins by removing inhibitory region
give ex of proprotein convertases
proinsulin is made as one inactive polypeptide
convertases remove middle section, then 2 remaining sections form active insulin
prevents premature signalling by insulin at the er
brings N and c terminus together
when stimulus = cuts proteins
middle acts as repressor
no specific motifs but some combos recognized better
makes disulfide bonds when passes through er
give ex of regulation by proteases
protein cleaved in golgi - upr
golgi signal usually hidden by chaperones
n terminus of protein = active transcription site
ATF6 is activated by convertase proteolysis in the Golgi
Regulation is by trafficking= BiP covers ER exit signal on ATF6 and proteases are only in Golgi
how does rab gtpase work
works same as ras
when active = anchored through 2 prenyl groups - anchored into membrane
describe rab proteins
Large subfamily of Ras-related proteins
Different organelles and vesicle types in secretory pathway have unique sets of Rab proteins
Switched “on” by GEF, “off” by GAP
Rab-GTP binds to a large number of Rab-effector proteins which mediate vesicle targeting
name rab functions
act at several steps in vesicle targetting
assist cargo selection and coat formation during vesicle budding
connect vesicle to motors on cytoskeleton for transport
tether vesicles to acceptor membrane - specificity
does many things, but other components = cofactors
Recognize cargo
transport, tethering, fusion
describe rab membrane anchor
Rabs have two prenyl lipid groups attached at their C-termini
describe rab in gdp bound state
In the GDP-bound state, Rab is soluble and not associated with the membrane
lipid is covered up by other proteins (GDI,GDF)
to hide them (prenyl groups)
describe rab in gtp bound state
In the GTP-bound state, the lipid modifications are exposed and anchor Rab to the membrane
theRab-effector proteins become attached to the membrane through Rab-GTP
sometimes has amphiphatic hexlix - stronger, tighter
describe rab activation
Specific GEF on membrane produces anchored, active Rab-GTP= GEF is linked to formation of vesicle coat
rab gtp works through effector proteins
describe how rab gtp works through effector proteins
attach vesicle to motor proteins
tether vesicle to target membrane
activate PI kinases and GEFs to make more Rab-GTP in clusters on acceptor membrane
describe 4 steps of vesicle rab cycle
- Vesicle Rab is activated by
GEF on donor membrane and packaged onto vesicles- some interact with cargo, assist uncoating, or attach to motors - Vesicle Rab-GTP binds specifically to tethers on acceptor membrane
- After fusion,GAPs on target membrane inactivate Rab
- Inactive vesicle Rab-GDP is recycled through cytosol to donor membrane
describe cytoskeleton
protein filaments - actin - and microtubules - tubulin - run through cytosol and provide structure to cells
vesicles travel with molecular models
connected to each other and anchored to organelles and pm (cortical skeleton)
usually localized in specific places, like in synapses for neurons, for microtubules = long so goes from nucleus to end of cell
describe actin filaments
shorter
often clusteres at pm
highly cross linked into network
motor = mysosins
