In most cells, cAMP inhibits cellular proliferation by blocking signaling through the ras-MAPK pathway. But in a subset of cell types cAMP functions as a potent mitogen that mediates the proliferative effect of certain endocrine hormones. The mechanism by which cAMP regulates cellular proliferation in these cells is unknown; but the phosphorylation dependent activator CREB has been implicated in this process. Recent work in our laboratory suggests, for example, that CREB functions importantly in endochondral bone development. The major hypothesis to be tested is that CREB induces proliferation of growth plate chondrocytes during by stimulating expression of cell cycle regulatory genes in response to cAMP. This hypothesis will be tested in the following aims: 1. We will test whether CREB family members are required for proliferation of endocrine target tissues by generating transgenic animals expressing a dominant negative CREB expression vector, designated A- CREB, in growth plate chondrocyte of endochondral bones. 2. We will test whether CREB activity is sufficient to promote proliferation of growth plan chrondrocytes during developing by preparing transgenic mice that express a constitutively active (Y134F) mutant CREB transgene under control of the rat collagen type II promoter. 3. We will employ gene profiling technology to identify cellular genes that are differentially expressed in chondrocytes of transgenic of transgenic mice expressing dominant negative (A-CREB) or constitutively active (Y134F) CREB polypeptides compared to wild type littermates. CREB target genes will be evaluated for their ability to rescue the proliferative defect in A-CREB transgenic chondrocytes.
