P1246990953cYLqg - PowerPoint PPT Presentation

Title: P1246990953cYLqg


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The secretory pathway of budding yeast
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Mossessova et al., Cell 2003, Miller et al.,
Cell 2003
C-site
B-site
LxxME DxE
A-site NPF
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How does protein folding influence COPII budding?
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The ABC transporter, Yor1p, as a model
folding/trafficking substrate.
lumen
cytoplasm
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Oligomycin sensitivity of Yor1p mutants
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An in vitro vesicle budding assay.
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Blue Native gel electrophoresis reveals
folding/assembly defects associated with Yor1
mutants.
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Genetic approaches to understanding Yor1p folding
and secretion
Multi-copy suppressor screen overexpress yeast
genomic fragments -- chaperones, folding/export
factors Extragenic suppressor screen
mutagenize yeast genome -- quality control
checkpoints, degradation machinery
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Screening the yeast genome for ER retention
defects.
(iv) confirm defects by assaying more substrates,
examining morphology etc.
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Understanding the regulation of ER export in the
context of protein folding and assembly.
biochemical characterization of
chaperone/cargo/coat interactions and
identification of proteins at ER exit sites
genetic selections to identify regulators of ER
export chemical screens to identify compounds
that overcome ER retention of misfolded proteins
(eg CFTR-related proteins)
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Acknowledgments
Columbia Justine Barry Marcus Lee
U Iowa Scott Moye-Rowley Dartmouth Charlie
Barlowe UC Berkeley Randy Schekman