The regulation of target gene expression by the nuclear receptor superfamily is modulated by both physiologic and pharmacological ligands and by the actions of various signal transduction pathways that result in either ligand-independent gene activation or down regulation of activity. The applicant has shown that this pathway-dependent crosstalk is mediated by association of nuclear receptors with complexes containing proteins which possess histone acetylase activity in the case of activation events, or with Nuclear receptor CoRepressors (N-CoR or SMRT) to mediate repression. He has demonstrated that the transcriptional activity of PPARgamma in response to treatment of cells with troglitazone is dependent upon an association with the receptor of a complex which contains several Nuclear receptor CoActivators (NcoAs) which include Creb-binding protein (CBP), CBP-interacting protein (pCIP), CBP associated factor (pCAF), and NcoA1. Conversely, treatment of cells with mitogens or expression of activated H-ras results in the subsequent recruitment to the PPARgamma receptor of a corepressor complex which includes the mammalian homologue of the yeast deacetylase RPD3, functionally mediated by a protein referred to as Silencing Mediator of Retinoid and Thyroid receptors (SMRT) rather than by the closely related N-CoR protein. In this application, the applicant proposes to test the hypothesis that differential recruitment of these coregulatory complexes represents the basis for combinatorial control of gene expression in response to PPARgamma ligands and thus regulates PPARgamma dependent processes, such as differentiation.
Three specific aims are proposed in the application. The first is to characterize the coactivator complexes required for transcriptional activation by PPARgamma, and to identify their specific roles in differentiation and thiazolidinedione activity in various tissues. The second involves an understanding of co-repressor activities as they pertain to PPARgamma function and the interaction of this regulatory mechanism with cellular signal transduction pathways. The third relates to an analysis of the role of leucine and charged amino acid domains in the regulation of PPARgamma and other nuclear receptors, allowing a further appreciation of the physical interactions required for the formation of a transcriptionally active complex. These studies will provide significant new insights into the regulation of PPARgamma activity and may suggest new therapeutic targets for increased insulin sensitivity in target tissues.
