Transcript Getting things where they need to go: Protein Targeting

Previously Bio308
Hypotheses for molecular basis of bipolar disorder
•Suggest problem lies in protein targeting
Proteins made in cytosol (cytosolic and membrane ones)
Sorting places proteins in membrane and in lumen of organelles
Importance of specific translocation
>50% of protein made on cytosolic ribosomes are not intended
to be used in the cytosol
Must cross between 1 and 3 membranes to reach final destination
Mis-localization can have drastic consequences—disease or death
How does the cell know where to place a protein?
Cellular ‘ZIP code’
Signal Sequences and Signal Patches
Signal sequences
Need a ‘routing code’
A section of the protein’s amino acid sequence is necessary and
sufficient to tell the cellular machinery where to place that protein
Targeting to the ER
If targeted to the ER where can a protein end up?
Main point of entry into the endomembrane system
TWO methods of targeting to ER
Minor pathway: Sec-dependent translocation
Identified first in bacterial genetic screens
Post translational
Post-translational translocation
Sec- dependent
Co-translational translocation
Major pathway: SRP-dependent translocation
First identified in in vitro experiments using canine microsomes
and wheat germ translation systems
Co-translational
CBI 12.3
Co-translational translocation
Important components from ER: SRP- receptor, TRAM
Sec61 complex (& BiP/Kar2-- sometimes)
Mammals: ER translocation involves “push”
Yeast: ER translocation involves “push” and “pull”
So the protein can now be in the ER-*Where in the ER
and then what happens?
ER proteins
ER
Where can a protein end up in the ER? How does it get there?
Lumenal proteins
Single transmembrane span proteins
Multipass transmembrane proteins
What category do our neurotransmitter and
neurotransmitter receptor fall in?
Getting out of the ER
Golgi enzyme involved in glycosylation
Neurotransmitter
receptor
ER
Lysosomal acid hydrolase
Now what?
Vesicular traffic
Secretory pathway: also method for delivering new PM proteins
ER to Golgi to trans-Golgi network
 then constitutive or regulated exocytosis
Constitutive and Regulated Exocytosis
Constitutive= constant, sometimes called ‘bulk flow’
Constitutive does not mean ‘un-regulated’
Regulated= needs additional signal to initiate fusion of vesicle with PM
Consequences of unregulated vesicular traffic
Mixing of organelle contents ( won’t function correctly)
Mislocalization of proteins ( won’t function correctly)
Inappropriate levels of secretion (too hi or too lo)
A Dead Cell
Vesicular traffic control
Our neurotransmitter receptor need to go ‘through’
5 cellular compartments before it gets to the post synaptic membrane
How does a vesicle ‘know’ what components it should contain?
How does it ‘know’ which membrane it should go to?
How does it fuse when it gets there?
Stages of vesicle traffic
3 Stages: Budding, targeting/docking and fusion
Donor
Donor
Target
Target
Content selection
What goes inside which vesicle?
Lumenal protein:
Transmembrane proteins:
Combination of cytosolic and
lumenal proteins determine
specific vesicle content
Budding
Fig 17-58
CBI 13.1 Clathrin
The SNARE hypothesis
V-SNARE
T-SNARE
Role of Rab proteins
retrograde
Fig 17-59
Synaptic vesicle fusion
VAMP
Syntaxin
SNAP 25
Rab3a
Synaptotagmin