PKC ISOZYMES AND INTESTINAL EPITHELIAL GROWTH CONTROL

Understanding of the signaling pathways that orchestrate the process of intestinal epithelial self-renewal is still incomplete. Based on increasing evidence for the role(s) of members of the protein kinase C (PKC) family of signal transduction molecules in control cell growth and differentiation, the long-term goals of the research proposed in this application are (a) to define the function(s) of individual PKC isozymes in regulation of the renewal process in the normal intestine, and (b) tp determine how alterations in PKC isozyme signaling pathways contribute to the development of intestinal disease. Our previous studies support the hypothesis that individual PKC isozyme(s) are key components of signaling pathway(s) that regulate intestinal epithelial cell growth/cell cycle progression, and that aberrations in these pathways contribute to the dysregulate cell growth seen in neoplastic disease and other conditions. To test this hypothesis, an integrated in vitro and in vivo approach will be used to address the following Specific Aims: (1) to investigate the ability of individual PKC isozyme(s) to control cell cycle progression in the IEC-18 intestinal crypt cell lines, using tetracycline-regulated machinery that are involved in PKC isozyme- mediated IEC-18 cell cycle arrest, using a variety of biochemical and molecular approaches, (3) to define the signaling events involved in PKC-mediated induction of the cyclin dependent kinase inhibitor p21/waf/cip1 in IEC-18 cells, by examining the MAPK pathway, the p53 growth suppressor protein and the Sp family of transcription factors, (4) to investigate the growth-regulatory functions of PKC alpha in the intestinal epithelium in vivo, using chimeric-transgenic mouse models over-expressing wild-type or dominant-negative PKC alpha in crypt and villus cells, and (5) to define the sequence of alterations in individual PKC isozyme expression and subcellular distribution during colon carcinogenesis, using the Min mouse and the dimethylhydrazine model of chemically-induced colon carcinogenesis in mice (and rats). These studies are expected to enhance our understanding of the role(s) of PKC isozyme(s) in intestinal epithelial cell growth control and tof their contribution to the development of colonic neoplasia.