REGULATION OF MITOGENIC CYTOKINE INDUCED APOPTOSIS

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DESCRIPTION (Adapted from Investigator's Abstract). Cell proliferation requires coordinated activation of genes involved in cell cycle progression as well as genes for maintaining cell viability. Without the latter, progression through the cell cycle will lead to apoptosis. The applicant has investigated the relationship between cell proliferation and apoptosis by activating cells with mitogenic cytokines and growth factors in the presence or absence of serum. When stimulated with cytokines or growth factors in the absence of serum, several cell types underwent apoptosis, a phenomenon referred to as cytokine-promoted apoptosis (CPA). Interestingly, in this system, serum induces expression of cell survival genes such as bcl-2 while mitogenic cytokines induce expression of pro-apoptotic genes including bax and c-myc in the absence of serum. By transfecting cells with an expression cDNA library, followed by double selection with CPA and neomycin, the applicant has identified a novel cell survival gene SRG-1. This gene is located on human chromosome 1 at 1p34-1p36.1, a location implicated in tumorigenesis in a number of cell lineages. SRG-1 encodes a putative protein of 154 amino acids. Stable transfection of SRG-1 to BAF/B03 cells confers resistance to CPA. SRG-1 also inhibits c-myc-driven apoptosis in serum-deprived fibroblasts. SRG-1 is highly expressed in some tumors and in activated T cells, but is not detected in most normal tissues, except liver and spleen. Based on these preliminary data, the applicant proposes to determine the mechanism underlying CPA and to characterize the role of SRG-1 in regulating CPA. The applicant hypothesizes that cytokines induce expression of genes like c-myc, which predispose cells to proliferation or undergo apoptosis. The ultimate outcome is determined by the presence or absence of second signals which activate cell survival genes such as Bcl-2 and SRG-1. The following aims are designed to test the hypothesis: 1) The applicant will establish the role of c-myc, its upstream Realtor E2F1 and a downstream target cdc25A in CPA. These genes will be modulated by transfection of the cDNA in anti-sense and dominant negative forms under the control of the ecdysone-inducible promoter. 2) The applicant will determine the role of SRG-1 in CPA by examining its expression pattern, cell survival functions, and interacting proteins. The completion of the proposed experiments may improve the understanding of the regulation of cellular homeostasis during inflammation, development, aging and tumorigenesis.