The long-term goal of my research is to elucidate the mechanisms that mediate cargo selection and export from the endoplasmic reticulum (ER). The cytosolic COPII coat complex is the basic sorting machine that mediates ER export. Selective low affinity interactions of COPII components with ER export signals on cargo) proteins support cargo selection and incorporation into COPII vesicles. We hypothesized that these low affinity interactions are transiently transformed into high avidity multivalent protein-lipid and protein-protein interactions at budding sites to enable ER export. The Sar1p GTPase may control this """"""""membrane transformation"""""""", through effector activation and membrane binding. Membrane remodeling may be essential to support the dynamic and localized nature of coat assembly and disassembly. Dynamic lipid re-modeling may also be required to enable localized membrane curvature and vesicle fission. In preliminary results, we demonstrate that local membrane re-modeling, through activation of phospholipase D and PI-kinases, may contribute to coat assembly and ER export. The involved enzymatic activities will be characterized in biochemical assays, and the temporal re-modeling of ER membranes, will be analyzed using morphological in vitro assays with fluorescent lipid-binding peptide reporters. In preliminary results, we also demonstrate that Sar1p interactions with re-modeled bilayers support vesicle fission. We will characterize the molecular basis for Sar1p and COPII Sec23/24 complex interactions with lipid bilayers. The role of coat-lipid interactions in vesicle formation, cargo selection and vesicle fission, will be studied using combined biochemical morphological and biophysical vitro and in vivo assays. The results may provide a general model to describe the mechanism and dynamics of membranes deformation, cargo selection and vesicle fission in vesicular traffic. ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK062318-03
Application #
7008160
Study Section
Cell Development and Function Integrated Review Group (CDF)
Program Officer
Haft, Carol R
Project Start
2004-03-01
Project End
2009-02-28
Budget Start
2006-03-01
Budget End
2007-02-28
Support Year
3
Fiscal Year
2006
Total Cost
$287,120
Indirect Cost
Name
University of Pittsburgh
Department
Physiology
Type
Schools of Medicine
DUNS #
004514360
City
Pittsburgh
State
PA
Country
United States
Zip Code
15213
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