The long term broad objective of this proposal is to uncover molecular mechanisms mediating cellular protein transport and sorting through the secretory pathway. Recent studies are revealing a central role for the cytoskeleton in the function and organization of the secretory pathway. Cytoskeletal regulation in the secretory pathway has been linked to cell transformation. Thus, this work will provide insight into cancer etiology and the invasive behavior of cancer cells, A signaling pathway has been characterized that connects proteins involved in COPI transport vesicle assembly at the Golgi with proteins that regulate the assembly of the cytoskeleton. This pathway depends on a cargo-protein-sensitive binding interaction between the coat protein, coatomer, and the GTP-binding protein Cdc42. Activation of this pathway leads to the recruitment of a specific actin-binding protein, mAbp1 to the Golgi. Both upstream (Cdc42) and downstream (mAbp1) components of this signaling pathway are involved in protein transport at the Golgi apparatus. Preliminary results link the coatomer/Cdc42-dependent signaling pathway to the recruitment of the molecular motor dynein to the membrane. It is hypothesized that a role for vesicle-coat dependent signaling to the cytoskeleton is to provide precise temporal regulation between the processes involved in transport vesicle assembly and cytoskeleton-mediated vesicle scission and translocation, As a first step toward proving this hypothesis, the following aims will be carried out:
Aim 1) To reconstitute coatomerdependent cytoskeletal regulation on liposomes, Aim 2) To define the domains of mAbpl that are important for its recruitment to the Golgi.
Aim 3) to determine how dynein recruitment responds to coatomer-Cdc42 signaling at the Golgi.
Aim 4) to determine which p24-family vesicle cargo proteins influence coatomer/Cdc42 signaling.
These aims will define the role of the coatomer/Cdc42 complex in cytoskeletal regulation, define the role of coatomer-dependent actin signaling in dynein recruitment and identify novel proteins that function in cytoskeletal regulation at the Golgi.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM068674-03
Application #
6930342
Study Section
Cell Development and Function Integrated Review Group (CDF)
Program Officer
Shapiro, Bert I
Project Start
2003-08-01
Project End
2007-07-31
Budget Start
2005-08-01
Budget End
2006-07-31
Support Year
3
Fiscal Year
2005
Total Cost
$245,219
Indirect Cost
Name
University of Iowa
Department
Physiology
Type
Schools of Medicine
DUNS #
062761671
City
Iowa City
State
IA
Country
United States
Zip Code
52242
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Hehnly, Heidi; Xu, Weidong; Chen, Ji-Long et al. (2010) Cdc42 regulates microtubule-dependent Golgi positioning. Traffic 11:1067-78
Hehnly, Heidi; Longhini, Katrina Marie; Chen, Ji-Long et al. (2009) Retrograde Shiga toxin trafficking is regulated by ARHGAP21 and Cdc42. Mol Biol Cell 20:4303-12
Hehnly, Heidi; Stamnes, Mark (2007) Regulating cytoskeleton-based vesicle motility. FEBS Lett 581:2112-8
Hehnly, Heidi; Sheff, David; Stamnes, Mark (2006) Shiga toxin facilitates its retrograde transport by modifying microtubule dynamics. Mol Biol Cell 17:4379-89
Pelish, Henry E; Peterson, Jeffrey R; Salvarezza, Susana B et al. (2006) Secramine inhibits Cdc42-dependent functions in cells and Cdc42 activation in vitro. Nat Chem Biol 2:39-46
Xu, Weidong; Stamnes, Mark (2006) The actin-depolymerizing factor homology and charged/helical domains of drebrin and mAbp1 direct membrane binding and localization via distinct interactions with actin. J Biol Chem 281:11826-33
Chen, Ji-Long; Fucini, Raymond V; Lacomis, Lynne et al. (2005) Coatomer-bound Cdc42 regulates dynein recruitment to COPI vesicles. J Cell Biol 169:383-9
Chen, Ji-Long; Xu, Weidong; Stamnes, Mark (2005) In vitro reconstitution of ARF-regulated cytoskeletal dynamics on Golgi membranes. Methods Enzymol 404:345-58
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