The PI proposes to determine and characterize the role of villin and its ligands (phosphatidylinositol 4,5- bisphosphate (PIP2) and actin) in epithelial cell motility. Active cell motility regulates many important intestinal epithelial cell functions, including: ion transport proteins via endocytosis and exocytosis, crucial for absorption of nutrients; intestinal restitution, important to maintain homeostasis in the presence of large osmotic and mechanical stress; the movement of cells along the crypt-villus axis; the invasion and propagation of enteropathogens; immune surveillance and inflammation; as well as neoplastic tumor cell dissemination and metastasis. Villin is an actin nucleating, capping, severing, and bundling protein. Villin binds and regulates two ligands that are known to regulate cell motility, phospholipase C-gamma1 (PLC-gamma1) as well as the substrate of the lipase namely, phosphatidylinositol 4,5-bisphosphate (PIP2). Recent studies with villin knock out mice have demonstrated that villin is necessary to regulate epithelial cell motility. In addition, villin shares sequence homology with other proteins of its family including gelsolin, which have been shown to regulate cell motility in vivo. Our working hypothesis is that villin's ability to regulate phosphoinositide-mediated signal transduction pathways and the actin cytoskeleton is important to epithelial cell physiology and pathophysiology involving changes in cell motility. To accomplish our overall study objectives we will characterize the villin-PIP2 interaction using the following approaches: reconstitution in vitro using recombinant villin proteins; endogenous villin expression in the intestinal epithelial cell line, Caco-2; and over expression of villin and enzymes that regulate intracellular PIP2 levels, in the villin null intestinal cell line, IEC-6 using a tetracycline-regulated system. To determine unequivocally the role of villin in the epithelial cell motility, we will use villin knock out mice. These studies promise the prospect of modifying motility for enhancement of normal physiology and for amelioration of disease. Inhibition of epithelial cell motility can be significant in several diseases, including inflammatory bowel disease, celiac disease, and colon cancer.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK065006-02
Application #
6849738
Study Section
General Medicine A Subcommittee 2 (GMA)
Program Officer
May, Michael K
Project Start
2004-04-01
Project End
2009-03-31
Budget Start
2005-04-01
Budget End
2006-03-31
Support Year
2
Fiscal Year
2005
Total Cost
$328,500
Indirect Cost
Name
University of Tennessee Health Science Center
Department
Physiology
Type
Schools of Medicine
DUNS #
941884009
City
Memphis
State
TN
Country
United States
Zip Code
38163
George, Sudeep P; Chen, Hongfeng; Conrad, Jacinta C et al. (2013) Regulation of directional cell migration by membrane-induced actin bundling. J Cell Sci 126:312-26
Wang, Y; George, S P; Srinivasan, K et al. (2012) Actin reorganization as the molecular basis for the regulation of apoptosis in gastrointestinal epithelial cells. Cell Death Differ 19:1514-24
Tomar, Alok; George, Sudeep P; Mathew, Sijo et al. (2009) Differential effects of lysophosphatidic acid and phosphatidylinositol 4,5-bisphosphate on actin dynamics by direct association with the actin-binding protein villin. J Biol Chem 284:35278-82
Khurana, Seema; Tomar, Alok; George, Sudeep P et al. (2008) Autotaxin and lysophosphatidic acid stimulate intestinal cell motility by redistribution of the actin modifying protein villin to the developing lamellipodia. Exp Cell Res 314:530-42
Khurana, Seema; George, Sudeep P (2008) Regulation of cell structure and function by actin-binding proteins: villin's perspective. FEBS Lett 582:2128-39
Wang, Yaohong; Srinivasan, Kamalakkannan; Siddiqui, Mohammad Rizwan et al. (2008) A novel role for villin in intestinal epithelial cell survival and homeostasis. J Biol Chem 283:9454-64
Mathew, Sijo; George, Sudeep P; Wang, Yaohong et al. (2008) Potential molecular mechanism for c-Src kinase-mediated regulation of intestinal cell migration. J Biol Chem 283:22709-22
George, Sudeep P; Wang, Yaohong; Mathew, Sijo et al. (2007) Dimerization and actin-bundling properties of villin and its role in the assembly of epithelial cell brush borders. J Biol Chem 282:26528-41
Wang, Yaohong; Tomar, Alok; George, Sudeep P et al. (2007) Obligatory role for phospholipase C-gamma(1) in villin-induced epithelial cell migration. Am J Physiol Cell Physiol 292:C1775-86
Tomar, Alok; George, Sudeep; Kansal, Pallavi et al. (2006) Interaction of phospholipase C-gamma1 with villin regulates epithelial cell migration. J Biol Chem 281:31972-86

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