The long term goal of my research program is to determine the molecular mechanisms by which extracellular signals regulate adipogenesis and the expression of adipocyte-specific genes. C/EBPa plays an essential role in development of adipocytes and the acquisition of insulin signaling in adipocytes. Given the importance of C/EBPa for adipocyte differentiation and metabolism, it is important to understand the molecular mechanisms by which CIEBPa transcription and activity are regulated. My laboratory has identified five sites of CIEBPa phosphorylation in vivo. Of these, T222 and T226 are phosphorylated by glycogen synthase kinase 3 (GSK3) and insulin stimulates dephosphorylation of these sites through inactivation of GSK3. During the course of these experiments, we discovered that inhibition of GSK3 blocks preadipocyte differentiation. Consistent with developmental effects of GSK3 being associated with signaling by Wnt, exposure of preadipocytes to Wnt blocked their ability to undergo adipogenesis. Based upon these findings, we hypothesize that inactivation of GSK3 by insulin and Wnt influences adipocyte gene expression and adipogenesis through regulation of C/EBP phosphorylation and activity.
The SPECIFIC AIMS of this grant application are to: 1) Determine the role of phosphorylation in regulation of C/EBPa activity by insulin. Experiments to investigate effects of phosphorylation of C/EBPa on transactivation and mitosis are proposed. The role of phosphorylation in regulating a subset of C/EBPa-dependent preadipocyte and adipocyte genes will be determined. 2) Determine the mechanism by which Wnts inhibits preadipocyte differentiation and the physiological significance of this pathway. Experiments will identify and investigate the regulation of Wnt-signaling and Wnt-regulatory proteins that act in adipocyte development. Experiments are proposed to determine whether repression of C/EBPa and PPARy gene expression by Wnt is mediated through b-catenin-TCF, Myc, GATAs, or whether inhibition of GSK3 by Wnt stimulates dephosphorylation and inactivation of C/EBPbeta. Understanding how insulin and Wnt signal through GSK3 and C/EBP transcription factors to regulate adipogenesis and adipocyte gene expression will provide important insight into the medical problems of obesity and type H diabetes, two major health risks in the United States.
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