""""""""This Section has focused on the role of Ca++ as a regulator of gene expression and cellular physiology. Using CAI as a tool, we have investigated Ca++-regulated molecular events. A novel gene, CAIR-1, was cloned from CAI conditioned cells, expressing a 74 kDa protein. Studies over the past year have investigated the signal transduction function of CAIR-1 and were initiated to identify a function for CAIR-1. We have demonstrated by cellular fractionation and immunohistochemistry that CAIR-1 is a cytosolic protein. It is expressed in vascular tissue and epithelial cells with no clear difference yet defined between tumor and normal cells; there is minimal expression in stroma. It is present in the epithelial cells of ovarian carcinoma specimens and stains their small vessels as well. A borderline ovarian cancer specimen also showed limited staining for CAIR-1. Pre- and post- CAI treatment specimens will be obtained in the current phase II CAI clinical trial and will be tested for changes in CAIR-1 expression. CAIR-1 function studies are being driven by putative functional domains in the protein. The PXXP regions suggest that CAIR-1 should bind SH3-containing proteins. Our hypothesis that CAIR-1 should function downstream of a calcium-regulated or calcium-associated pathways led to the identification of phospholipase C-gamma (PLC-g) as a putative partner protein. Coimmunoprecipitation studies indicate that CAIR-1 binds to PLC-g under basal conditions and is dissociated when cells are stimulated with epidermal growth factor. A manuscript describing the cloning and initial characterization of CAIR-1 has been submitted. A second approach to study the effect of Ca++ on gene regulation has used CAI as a tool to differentiate between a requirement for increased intracellular Ca++ and its source: intracellular release v. influx. Our recent collaboration with Dr. Kevin Gardner has identified a CAI- and calcium influx-sensitive event regulating the HIV long terminal repeat. This was demonstrated to have its effect through NFKB regulation. This further supports our observations that transmembrane calcium influx is a regulatory event for gene transcription. A final direction has been the investigation of extracellular calcium as a regulatory signal in invasion and metastasis. We have now demonstrated the presence of a functional calcium sensing receptor in primary culture human umbilical vein endothelial cells. Our current work focuses on demonstration of function for this receptor, validating our hypothesis that the extracellular calcium milieu is also regulatory.""""""""
Showing the most recent 10 out of 22 publications
