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， which哪一个to apply应用your emerging新兴understanding理解of molecular分子mechanisms机制of nuclear核receptor受体actions行动and 在定义transrepression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辅阻遏物复合物作为转录检查点控制配体依赖的 核受体对基因表达的调控及信号依赖性转录活性 驱动基因表达的炎症程序的因素。我们将更好地定义 N-CoR辅阻遏物复合物的TBLi、TBLRi和GPS2组分在脑缺血再灌注中的作用机制 调节APi/NF-kB靶基因，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复合物的假设 调节导致胰岛素抵抗的炎症反应，是抗糖尿病作用的靶点 PPARy激动剂。这些研究将与单位i和3合作进行，使用小鼠 用N-CoR "/"或SMRT "/"胎肝造血祖细胞重建。具体目标2将 研究TBLi/TBLRi交换复合物受信号特异性 TBLRi/TBLi的磷酸化。我们将研究辅阻遏物复合物的蛋白激酶调控 AP-i和NF-kB靶基因的消除，以及这些事件在PPARy介导的 正调控基因和炎症反应基因的反式阻遏。第3章将 探索GPS2和KIAAiySy在JNK依赖性基因激活/抑制事件中的作用，并检测 假设GPS2是正常胰岛素敏感性所必需的，基于JNK表达 和活性在饮食诱导的肥胖模型中持续升高，并且AP-1活性是组成性的， 在N-CoR +/-巨噬细胞中的基因靶点子集上增加。这些研究将结合使用 单细胞核显微注射siRNA，一种超灵敏的多重RNA定量方法 （RASL）和ChIP-DASL来定义GPS2在炎症反应的信号依赖性活化中的作用 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.