Trafficking 1 from ER to Golgi

1

• Summary
• Rough ER and smooth ER
• Signal hypothesis, translocation into ER
• Single-span and multi-span membrane proteins
• Glycosylation
• Protein folding
• Lipid synthesis

2
Lecture 8 Vesicular trafficking from ER to Golgi
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Endocytic and biosynthetic-secretory pathways
(Ten or more chemically distinct,
membrane-enclosed compartments)
Transport vesicles
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The biosynthetic-secretory and endocytic pathways
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Various types of coated vesicles
Golgi apparatus Plasma
ER and Golgi Cisternae
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Assembly of a clathrin coat
Coated pits and vesicles on the cytosolic
surface of membranes
36 triskelions 12 pentagons 6 hexagons
triskelion
Inner layer binds adaptins
Freeze-etch
7
Adaptin binds to cargo receptor and clathrin
triskelion
Dynamin pinches off the bud
Auxillin-activated ATPase is required To remove
the clathrin coat
Four types of adaptins
Vesicles can have different shapes
8
Dynamin pinches of the vesicles
GTPase
Shibire mutant has coated pits but no budding
off of synaptic vesicles
9
Assembly and disassembly of coat by GTPases
GTPase works like a timer And cause disassembly
shortly After the budding is completed
Coat-recruitment GTPases
ARF proteins COPIclathrin Sar1 protein COPII
GTP causes Sar1 to Bind to membrane
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Guidance of vesicular transport SNAREs
specificity and fusion Rab GTPases initial
docking and tethering of vesicles to target
membranes and matching of v- and t- SNAREs
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SNARE proteins guide vesicular transport
20 SNAREs, v-SNAREs, t-SNAREs
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SNAREs specify compartment identity and control
specificity
4 a helices in trans-SNARE complexes
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Rab proteins ensure the specificity of vesicle
docking
gt30 Rabs
On cytosolic surface
C-terminal regions are variable Bind to other
proteins, including GEFs
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After docking
SNAREs may mediate membrane fusion
SNARE complex
15
The entry of enveloped viruses into cells
HIV
Similar to SNAREs
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Proteins leave the ER in COPII-coated transport
vesicles
ER exit sites (no ribosomes)
Selective process
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Only properly folded and assembled proteins can
leave the ER
Chaperones cover up exit signals
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Homotypic membrane fusion
to form vesicular tubular clusters
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Vesicular tubular clusters
Lacks many of the ER proteins
Short-lived
COPI-coated
Retrograde transport
carry back the ER resident proteins that leaked
out
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ER retrieval signals KKXX in ER membrane
proteins, KDEL sequence in soluble ER resident
proteins
pH controls affinity of KDEL receptors
Membrane proteins in Golgi and ER have shorter TM
domains (15 aa) Cholesterol
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Ordered series of Golgi compartments
Cisternae, tubular connections
Plant cell
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MT is required to localize near the cell nucleus
close to the centrosome (in animal cells)
Plant cells
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Two main classes of N-linked glycosilation
core
complex oligosaccharides
high-mannose oligosaccharides
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Oligosaccharide processing in the ER and the Golgi
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Why glycosylation?
Folding Transport Stability Recognition Regulator
y roles(Notch)
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Functional compartmentalization
Histochemical stains biochemical Compartmentaliza
tion of the Golgi
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Transport through the Golgi may occur by
vesicular transport or cisternal maturation (not
mutually exclusive)
Collagen rods Scales in algae
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• Summary
• Vesicular transport, biosynthetic-secretory and
• endocytic pathways
• 2. Coated vesicles
• Coat assembly and disassembly, budding, dynamin,
• coat-recruitment GTPases
• Targeting and fusion by Rab GTPases, SNAREs
• ER to Golgi COPII, folding, fusion (cluster),
  retrograde
• Golgi apparatus structure and polarity
• Continuation of glycosylation
• Compartmentalization of Golgi cisternae
• By now we have introduced gated transport,
  transmembrane
• transport and vesicular transport.