Redox-mediated p300 regulation of hepatocyte NF-kB

描述（由申请人提供）：细胞氧化还原状态调节所选转录因子（如NF-κ B）的活性。氧化还原依赖性NF-κ B活化的顺式和反式调节的传统一级范例已被广泛探索。然而，现在清楚的是，诱导型基因转录也需要跨调控区的染色质结构重组。研究表明，转录因子如NF-κ B与其靶位点的结合需要具有组蛋白乙酰转移酶（HAT）活性的转录辅激活因子的募集。这些辅激活蛋白反过来又受其磷酸化状态的调节。在氧化应激的特定情况下，很少有人知道的高级表观遗传机制，涉及氧化还原敏感的染色质重塑和组蛋白修饰NF-κ B依赖的转录。在白细胞介素-1 β（IL-1 β）刺激的CCL9.1小鼠肝细胞模型中，我们已经证明，过氧化物介导的氧化应激通过诱导组蛋白乙酰化和染色质重塑来增强诱导型一氧化氮合酶（iNOS）的转录，而组蛋白乙酰化和染色质重塑原本是沉默的NF-κ B结合位点（nt-114）增强NF-κ B结合。这种表观遗传现象不发生与IL-1 β或过氧化物单独。以前的研究还没有调查在肝细胞iNOS启动子的NF-κ B激活的氧化还原依赖性染色质重塑。在这个R21应用中，使用肝细胞iNOS作为氧化还原敏感性基因表达的模型，我们建议表征氧化还原依赖性转录共激活因子，其通过染色质重塑和组蛋白超乙酰化增强NF-κ B激活。这种共激活蛋白及其功能相关的氧化还原介导的染色质重塑必须进行进一步的研究之前，其特征。初步研究将集中在p300转录共激活因子/HAT蛋白。鉴于这一实验性项目的新颖性和高风险性，我们认为R21供资机制特别相关。

DESCRIPTION (provided by applicant): Cellular redox state regulates activity of selected transcription factors, such as NF-kB. The traditional first-order paradigms of cis- and trans- regulation of redox-dependent NF-kB activation have been extensively explored. However, it is now clear that inducible gene transcription also requires reorganization of chromatin structure across regulatory regions. Studies demonstrate that binding of transcription factors, such as NF-kB, to their target sites requires recruitment of transcriptional co-activator(s) with histone acetyltransferase (HAT) activity. These co-activator proteins are, in turn, regulated by their phosphorylation state. In the specific instance of oxidative stress, exceedingly little is known of the higher-order epigenetic mechanisms involving redox-sensitive chromatin remodeling and histone modification in NF-kB dependent transcription. In a model of interleukin-1beta (IL-1beta) stimulated CCL9.1 murine hepatocytes, we have demonstrated that peroxide-mediated oxidative stress enhances inducible nitric oxide synthase (iNOS) transcription by inducing histone hyperacetylation and chromatin remodeling at an otherwise silent NF-kB binding site (nt -114) to enhance NF-kB binding. This epigenetic phenomenon does not occur with IL-1beta or peroxide alone. Previous studies have not investigated redox-dependent chromatin remodeling in NF-kB activation at the hepatocyte iNOS promoter. In this R21 application, using hepatocyte iNOS as a model of redox-sensitive gene expression, we propose to characterize the redox-dependent transcriptional co-activator which enhances NF-kB activation through chromatin remodeling and histone hyperacetylation. This co-activator protein and its functional relevance to redox-mediated chromatin remodeling must be characterized before additional studies can proceed. Initial studies will focus upon the p300 transcriptional co-activator/HAT protein. Given the novel and high risk nature of this experimental venture, we believe the R21 funding mechanism to be particularly relevant.