Transcript The Exocytic/Lysosomal Transport Pathway

Vesicular Trafficking
Movement From the ER Through the
Golgi
Getting to the Golgi
Anterograde and Retrograde Traffic
Genetic Analysis of ER-Golgi Transport
General Principles
• Cargo selection
– signals (ie amino acid sequences)
• Vesicle formation (budding) from donor
compartment
• Vesicle/Cargo targeting and fusion to acceptor
compartment
• Most regulation is mediated by cargo- and
target-compartment-specific small GTPases
• Anterograde transport
– COPII apparatus
– protein complex mediating cargo selection and
assembly of budding components from ER
• Retrograde transport
– COPI
– protein complex mediating transfer of vesicles back to
ER
Retrograde Transport Gives Golgi
Compartments Dynamic Identity
Cargo Selection: Export Targeting Sequences
• Y xnDxE – tyrosine-diacidic domain
• Found in cytoplasmic tail of TM proteins
Cargo Selection: Just Getting the Right Stuff
• Exclusion of ER resident proteins
– not well understood
– fidelity factors (genes)
• mutations in some genes cause increased excretion of ER
resident proteins
– Retrieval
• COPI vesicles retrieve ER proteins and return them
• KDEL signal on lumenal proteins
• KKxx-COOH on TM proteins (dilysine motif)
COPII Vesicular Coat Assembly & Budding
• COPII coat
formation
regulated by
sar1 GTPase
• sar1 activates
cargo receptors
– TM proteins
recognizing
export signals
• sar1 recruits
sec23/24
coatamer
proteins
• sec23/24 in turn
recruit sec31/13
coat proteins
• COPII vesicles
form and bud
• Once apart from ER, coat proteins are triggered to release by
hydrolysis of GTP by sar1
Vesicular Delivery
• Rab-GTPases
• Recruits tether
and fusion
proteins into
vesicles
• These form a
targeting complex
recognized by a
docking complex
• targeting
– v-tethers – p115
– vSNAREs –
synaptobrevin
• docking
– t-tethers –
GRASP65
– tSNAREs –
syntaxin 1
Tethering and Fusion Mediated by SNAREs
(SNAP Receptors)
• SNAREs aggregated by Rab-GTPase (vSNAREs)
• Vesicle buds
• SNAP25-NSF interact with vesicle to dis-aggregate and prime
vSNAREs for interaction with tSNAREs
COPI
Vesicles
Recycle ER
Resident
Proteins
• COPI Vesicles
– Arf family of small GTPases
– Recruit/activate coatomer (6 cytosolic proteins)
– Recruit ER targeting SNAREs
Retrieval of ER
Proteins with KDEL
Signal by COPI
Vesicles
Cycle of Vesicular Transport
Cargo Sorting by the Trans-Golgi Network (TGN)
• Specific coat proteins direct TGN vesicles to targets
Lysosomal/Endosomal Targeting
• Mannose-6-phosphate
receptor targeting to
lysosome/endosome
• MPR cytoplasmic tail
(C-term) has multiple
signals for multiple
routing possibilities
Lysosomal
Targeting of
Prohydrolase by
M6P and MPRs
•Mannose-6-Phosphate Receptors
•Cation-independent (CI)
•Cation dependent (CD)
Cytoplasmic Tails of MPRs Allow Sorting to
Lysosomes vs Other Routes
Assembly of Clathrin on Lysosomal/Endosomal
Destined TGN Vesicles
Directed Translocation of Secretory Vesicles
• apical targeting in
caveolin coated, lipidraft vesicles
• basolateral targeting
signals
– direct protein to this
membrane
• constitutive basolateral
targeting
– subsequent sorting of
apical proteins
• untargeted vesicles
– sorting to redistribute
surface proteins
Proteolytic Processing of
Pro-proteins in Secretory
Vesicles
• Cleavage of proinsulin in
secretory vesicles of
pancreas cells
Secretory Vesicles/Granules
• Material in
secretory vesicle
aggregates due
to decreased pH
and divalent
cations
• Membrane
recycling to TGN
directly or via
endosome
reduced vesicle
size
Regulated Secretion
• Fusion of the vesicle and plasma membrane is
regulated by various means
– receptor-ligand interaction and second messenger
signaling
– depolarization of membrane
– etc…