The overall goal of this grant is to understand why polyamines are essential to the migration of GI mucosal cells during the early mucosal restitution component of healing. Experiments supported during the past 15 years this project has been active have shown that: 1) polyamines are essential to the healing of gastric and duodenal mucosa, 2) polyamines are required for mucosal restitution which depends on cell migration, 3) polyamines are essential to cell migration, 4) polyamines are necessary for the normal functioning of several components of the cytoskeleton involved in cell migration, 5) migration of IEC-6 cells is regulated by the Rho GTPases, and 6) polyamines influence the levels and activities of the Rho GTPases. The first set of specific aims will determine the effect of polyamines on the Rho family of GTPases during the migration of IEC-6 cells. IEC-6 cells stably transfected with constitutively active and dominant negative RhoA, Rac1 and Cdc42 will be used to determine their hierarchy of activation and where the polyamines interact. Preliminary data indicate that both RhoA and Raci are essential for cell migration but that only Rac1 activity is sufficient for migration. The second set of specific aims will determine the role of polyamines in integrin signaling by examining ccli attachment and spreading. The same clones and polyamine depletion will be used to examine the roles of Rho, Rac and Cdc42. Polyamine depletion will be used to determine the role of polyamines in the distribution and phosphorylation of proteins involved in the formation of focal adhesions and in cell spreading. In the third set of specific aims a highly specific polyamine antibody will be used to determine the cellular localization of polyamines during the processes of spreading and migration and to determine the colocalization of polyamines with cytoskeletal proteins and those regulating cytoskeletal transformations. These studies will provide the first description of the involvement of the Rho-GTPases in epithelial cell migration and spreading and how the activities of these proteins are regulated by polyamines. They also will provide the first studies of polyamine localization during dynamic cell processes and define the mechanisms by which polyamines regulate those processes. This work will increase our understanding of cell function basic to many clinical conditions in which cell migration is a major component. These include peptic ulcer disease, cancer, and inflammatory bowel disease.
| Ray, Ramesh M; Johnson, Leonard R (2014) Regulation of intestinal mucosal growth by amino acids. Amino Acids 46:565-73 |
| Ray, Ramesh M; Li, Chunying; Bhattacharya, Sujoy et al. (2012) Spermine, a molecular switch regulating EGFR, integrin ýý3, Src, and FAK scaffolding. Cell Signal 24:931-42 |
| Ray, Ramesh M; Viar, Mary Jane; Johnson, Leonard R (2012) Amino acids regulate expression of antizyme-1 to modulate ornithine decarboxylase activity. J Biol Chem 287:3674-90 |
| Ray, Ramesh M; Bavaria, Mitulkumar N; Bhattacharya, Sujoy et al. (2011) Activation of Dbl restores migration in polyamine-depleted intestinal epithelial cells via Rho-GTPases. Am J Physiol Gastrointest Liver Physiol 300:G988-97 |
| Bavaria, Mitulkumar N; Ray, Ramesh M; Johnson, Leonard R (2011) The phosphorylation state of MRLC is polyamine dependent in intestinal epithelial cells. Am J Physiol Cell Physiol 300:C164-75 |
| Elias, Bertha C; Bhattacharya, Sujoy; Ray, Ramesh M et al. (2010) Polyamine-dependent activation of Rac1 is stimulated by focal adhesion-mediated Tiam1 activation. Cell Adh Migr 4:419-30 |
| Ray, Ramesh M; Bhattacharya, Sujoy; Johnson, Leonard R (2007) EGFR plays a pivotal role in the regulation of polyamine-dependent apoptosis in intestinal epithelial cells. Cell Signal 19:2519-27 |
| Ray, Ramesh M; Guo, Huazhang; Patel, Minesh et al. (2007) Role of myosin regulatory light chain and Rac1 in the migration of polyamine-depleted intestinal epithelial cells. Am J Physiol Gastrointest Liver Physiol 292:G983-95 |
| Ray, Ramesh M; Vaidya, Rajiv J; Johnson, Leonard R (2007) MEK/ERK regulates adherens junctions and migration through Rac1. Cell Motil Cytoskeleton 64:143-56 |
| Bhattacharya, Sujoy; Ray, Ramesh M; Johnson, Leonard R (2006) Integrin beta3-mediated Src activation regulates apoptosis in IEC-6 cells via Akt and STAT3. Biochem J 397:437-47 |
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