Functional aspects of Adenomatous polyposis coli protein dimerization and posttranslational regulation

The Adenomatous Polyposis Coli (APC) tumor suppressor gene is responsible for an inherited predisposition to colon cancer through the development of multiple colorectal adenomas. Mutations in APC are also found in the majority of sporadic colorectal cancers, indicating that APC function is critical to maintenance of cell growth and differentiation in colonic epithelium. Soon after APC was identified, it was demonstrated that APC protein could form stable, coiled-coil homodimers via a predicted alpha-helical region within the amino terminus of the protein. Because this region of APC is predicted to be preserved in most mutant forms of the protein, it is likely that homodimer formation influences the function of both wild-type and mutant APC protein species. Using several approaches, I examined structural and biological aspects of the coiled-coil APC homodimerization domain. In a collaborative effort with the laboratory of Tom Alber at the University of California, Berkeley, I functionally tested a peptide probe that was designed to dimerize preferentially with the coiled-coil homodimerization domain of wild-type APC protein. The strategic design of peptide probes for APC protein provided the rationale and experimental tools for subsequent investigation into the effects of APC homodimer disruption in vivo. When expressed in cultured cells, these peptides interacted with endogenous wild-type APC protein in a manner that reflected their in vitro affinities for the wild-type APC dimerization domain; however, a preliminary survey revealed no clear effects of peptide expression upon known functions of APC protein. In addition to studying APC dimerization, I examined potential regulatory relationships between APC, beta-catenin and the ubiquitin-proteasome pathway. These experimental findings agree with published reports that beta-catenin is a substrate for the ubiquitin-proteasome pathway in vivo. Half-life analyses also reveal that wild-type APC protein is extremely stable in vivo, although truncated, mutant APC protein can be regulated by the ubiquitin-proteasome pathway.