In the next grant period we will focus on three of the seven mitogen- inducible TIS primary response genes we previously cloned. TIS10 encodes a second form of the enzyme prostaglandin synthase (PGS). TIS10/PGS2 is induced in a wide variety of cells, by many different stimuli, and is globally repressed by glucocorticoids. Regulation of TIS10/PGS2 gene expression is likely to play a major role in inflammation, reproduction, hemostasis, and wound healing. We will elucidate the mechanisms by which oncogenes, growth factors, steroid hormones, cytokines, and other signals regulate TIS10/PGS2 gene expression. We will use transfection experiments with reporter constructs, site-directed promoter mutation, DNA-affinity purification procedures, and genetic selections to (i) identify the cis- acting elements of the TIS10/PGS2 gene and the trans-acting factors that regulate TIS10/PGS2 expression and (ii) identity and characterize signal transduction pathways modulating TIS10/PGS2 transcriptional responses to extracellular signals. We think it likely that aberrant regulation of the TIS10/PGS2 gene contributes to pathophysiological processes that include arthritis, bone resorption, cardiovascular disease, atherosclerosis, immunosuppression, and colon cancer. We will determine the role of TIS10,PGS2 gene expression in normal and pathophysiological prostanoid production. We will use antisense oligonucleotide procedures to distinguish the in vivo roles of PGS1 E.C.1.14.99.1, the previously described PGS gene, and TIS10/PGS2. We will also characterize the phenotype of mice with TIS10/PGS2 gene disruptions. The TIS11 primary response gene is a member of a three gene family whose proteins have two copies of a cys-his finger. TIS11 and TlS11b proteins heterodimerize, and are phosphorylated following mitogen stimulation. Drosophila and yeast have TIS11 homologues that conserve both the cys-his repeat and the spacing between them. Existence of a gene family encoding proteins with a common motif, strong conservation of this motif from yeast to man, a wide range of inducibility, rapid and transient expression, and ligand-induced post-translational modification all suggest the TIS11 proteins have important roles. We will use biochemical and genetic approaches to identify the function(s) of the TIS11 proteins. The TIS21 primary response gene mRNA and protein are rapidly and transiently induced in many cell types. The TIS21 gene and protein share substantial similarity to the BTG1 gene and protein. BTG1 was identified because of its proximity to a chronic lymphocytic leukemia breakpoint. The stringent regulation, the short time window for the presence of TIS21 protein, its widespread inducibility, and the similarities of TIS21 and BTG1 suggest that the TIS21 protein also plays an important role in ligand-induced biological responses. We will use biochemical and genetic approaches to identify the function(s) of TIS21 protein.
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