The growth of mammalian cells is regulated by polypeptide growth factors. Understanding how such factors induce cell growth is an important step toward understanding the mechanisms controlling cell proliferation and neoplasia. Quiescent mouse fibroblasts can be stimulated to re-enter the cell cycle at the G1 phase by the administration of serum or purified growth factors. The interaction of growth factors with their receptors at the plasma membrane results in a signal transmitted to the nucleus, where transcriptional activation of a specific set of genes rapidly occurs. Among several genes known to be activated in this manner are the proto-oncogenes c-fos and c- myc. Accumulating evidence suggests that specific gene activation is a necessary step in growth factor-induced cell proliferation. Described in the preliminary studies section of this proposal is the isolation of cDNA clones corresponding to 10 previously unknown genes that are transcriptionally activated within minutes by platelet-derived growth factor, a potent mitogen in serum. Regulation of these genes occurs on transcriptional and post- transcriptional levels and is coordinate with the regulation of the proto-oncogene c-fos or c-myc. Among these """"""""immediate-early"""""""" genes, so named by analogy to the genetic program for viral development, are those anticipated to be key mediators of the proliferative response. Understanding the overall nature of these genes as a class and the biological activities of their gene products is the goal of this proposed research. The expression of these growth-related, immediate-early genes will be examined during growth, development, and differentiation in the mouse and in cell culture models. Five of these genes have been selected for further study. Their full-length cDNA sequences will be determined and their protein product primary structures will be deduced. Antisera against the protein products will be prepared; the protein products will be identified in cultured cells and in relevant tissues, and their subcellular locations will be defined. One of these genes will then be selected for further focus; the gene product will be purified and its biochemical activities analyzed. The effects of constitutive expression of this gene, inactivation by anti-sense RNA, and expression during phases of the cell cycle when it is usually silent will be investigated using a number of recombinant constructs.
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