Functional Char. of N-CoR/SMRT Corepressor Complexes in Adipocytes & Macrophages

巨噬细胞和胰岛素抵抗之间令人惊讶的关系提供了一个有希望的接口 你对核受体作用的分子机制的新理解和 在定义反压抑的基本策略方面的最新进展。在项目2中，我们将重点关注 NCoR/SMRT辅阻遏子复合体作为转录检查点控制配体依赖 核受体对基因表达的调控与信号依赖转录活性 驱动基因表达炎症程序的因素。我们将更好地定义分子 N-COR辅阻遏子复合体中TbLi、TbLRi和GPS2组分的作用机制 APi/NF-kB靶基因的调控，我们将使用全基因组定位分析(GWLA)来研究 这些蛋白在巨噬细胞和脂肪细胞基因表达的正负调控中的作用。 提出了三个具体目标。具体目标I将检验N-COR/SMRT复合体的假设 调节导致胰岛素抵抗并成为抗糖尿病作用靶点的炎症反应 PPARy激动剂。这些研究将与单元I和单元3合作，使用小鼠 用N-COR‘/’或SMRT/‘胎肝造血祖细胞重组。特定目标2将 探讨TbLi/TbLRi交换复合体受信号特异性调控的假说 TbLRi/TbLi的磷酸化。我们将研究核心阻滞剂复合体的蛋白激酶调控。 AP-I和NF-kB靶基因的取消以及这些事件在PPARy介导的激活中的作用 炎症反应基因的正调控基因和转录抑制。具体目标3将 探索GPS2和KIAAiySy在JNK依赖的基因激活/抑制事件中的作用，并测试 基于观察到JNK-表达是正常胰岛素敏感性所必需的GPS2的假设 在饮食诱导的肥胖模型中，AP-I活性持续升高，而且AP-I活性是结构性的 在N-COR“/‘巨噬细胞中增加了基因靶点的子集。这些研究将利用 单细胞核微量注射siRNAs--一种超灵敏的多重RNA定量方法 (RASL)和ChlP-DASL确定GPS2在信号依赖的炎症反应激活中的作用 基因。

The surprising relationship between macrophages and insulin resistance provides a promising interfacein which to applyour emerging understanding ofthe molecularmechanismsof nuclear receptor actions and recent progress in defining the underlying strategies oftransrepression. In Project 2, we will focus on NCoR/SMRT corepressor complexes as transcriptional checkpointscontrolling both ligand-dependent regulation ofgene expression by nuclear receptors and the activities of signal-dependent transcription factors that drive inflammatory programs of gene expression.We will better define the molecular mechanisms and roles ofthe TBLi,TBLRi and GPS2 componentsof N-CoR corepressor complexesin the regulation ofAPi/NF-kB target genes and we willuse genome-wide location analyses (GWLA) to investigate the roles ofthese proteins in positive and negative regulation of macrophage and adipocyte geneexpression. Three Specific Aims are proposed. Specific Aim i will test the hypothesis that N-CoR/SMRTcomplexes regulate inflammatory responses that contribute to insulin resistance and are targets ofanti-diabetic actions of PPARy agonists. These studies will be performed in collaboration with Units i and 3 using mice reconstituted with N-CoR'/'or SMRT/' fetal liver hematopoieticprogenitor cells. Specific Aim 2 will investigate the hypothesis that the TBLi/TBLRi exchangecomplexis regulated by signal-specific phosphorylation ofTBLRi/TBLi. We will investigate the protein kinase control of corepressorcomplex dismissal from AP-i and NF-kB target genes, and the role ofthese events in PPARy-mediated activationof positively regulated genes and transrepression ofinflammatory response genes. Specific Aim 3 will explore the role ofGPS2 and KIAAiySy in JNK-dependentgene activation/repression events and to test the hypothesis that GPS2is required for normal insulin sensitivity based on observations that JNK-expression and activityare consistently elevated in diet-induced obesity models and that AP-i activity is constitutively increased on a subset of genetargets in N-CoR"/' macrophages. These studies will utilize a combinationof single cell nuclear microinjectionof siRNAs, an ultra-sensitive, multiplexed RNAquantificationmethod (RASL) and ChlP-DASL to define roles of GPS2 in signal-dependent activation of inflammatory response genes.