We propose to study the regulation of the protein kinase C alpha (PKCalpha) promoter by two transcriptional regulators that play opposite roles during colon carcinogenesis, beta-catenin and retinoic acid receptor (RAR). PKCalpha functions as a tumor suppressor gene in colorectal cancer cells and its downregulation is associated with loss of the adenomatous polyposis coli (APC) tumor suppressor gene during the earliest stages of colorectal carcinogenesis. APC loss results in the accumulation of beta- catenin/Tcf-4 transcriptional complexes that regulate genes that are likely to play key roles in colorectal cancer development. Retinoids, which are ligands for the RAR family of transcription factors, induce PKCalpha expression and reverse the malignant phenotype of cancer cell lines, and prevent carcinogen-induced colon neoplasms in animal models. Thus, the regulation of PKCalpha expression by beta-catenin/Tcf-4 and RAR is of tremendous interest for understanding processes important in colorectal carcinogenesis. We cloned the murine PKC alpha promoter, which contains a functional response element for retinoic acid receptor as well as two putative T-cell factor (Tcf) response elements (TcfRE1 and TcfRE2). These observations provide the foundation of our CENTRAL HYPOTHESIS: PKCalpha is a transcriptional target of Tcf-4/beta-catenin and RAR during colon carcinogenesis.
Our SPECIFIC AIMS are to demonstrate that: 1) Tcf-4/beta-catenin regulates transactivation of the PKCalpha promoter through TcfRE1 and TcfRE2; 2) beta-catenin and RAR interact to regulate transactivation of the PKCalpha promoter; and 3) PKCalpha expression and parameters of malignant transformation are influenced by manipulation of beta-catenin stability and by ligands of RAR in vivo and in vitro. Our preliminary data strongly support our central hypothesis. Experiments proposed in this application will provide new insights into how beta-catenin and RAR interact to regulate expression of a tumor suppressor gene and may lead to novel chemotherapeutic and chemopreventative strategies that target these interactions.
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