Glucocorticoid Regulation of NF-kB Function by O-GlcNAc

描述（由申请人提供）：NF-κ B诱导的促炎基因活化可在激素结合后被糖皮质激素受体完全抑制。然而，将糖皮质激素信号传递到NF-κ B通路的关键辅因子和机制仍然难以捉摸。我们最近将O-GlcNAc转移酶（OGT）定义为基因阻遏中的非典型介质。因此，我们的第一个具体目标是探索这种酶作为糖皮质激素受体的候选辅阻遏物在肾上腺皮质激素反应中的作用。我们将通过生物化学方法测试OGT和糖皮质激素受体之间的物理相互作用，并通过基于细胞的基因报告基因测定来检查它们的功能相互作用。使用RNA干扰，我们将评估OGT在调节内源性促炎基因中的可能作用。与OGT不同的是，人们对相对酶O-Incase（OGN）对转录的影响知之甚少。因此，我们的第二个具体目标是辨别这两种相互作用的酶如何促进促炎基因网络的激活或抑制。使用染色质免疫沉淀法，我们将研究OGT和OGN在活性或抑制性促炎基因启动子的动力学。此外，我们将寻求通过生物化学方法确定这两种酶在转录装置中的上游调控因子和下游靶点。本研究旨在阐明O-GlcNAc调控转录的精确机制。由于糖皮质激素目前用于治疗炎症和癌症，通过O-GIcNAc了解它们的作用为治疗和药物设计提供了新的方法。

DESCRIPTION (provided by applicant): NF-kappaB-induced activation of proinflammatory genes can be completely repressed by the glucocorticoid receptor upon hormone binding. However, critical cofactors and mechanism that transmit the glucocorticoid signal into the NF-kappaB pathway remain elusive. We recently defined O-GIcNAc transferase (OGT) as an atypical mediator in gene repression. Hence, our first specific aim is to explore the role of this enzyme as a candidate corepressor for the glucocorticoid receptor in antiinflammatory responses. We will test a physical interaction between OGT and the glucocorticoid receptor by biochemical approaches and will examine their functional interaction by cell-based gene reporter assays. Using RNA interference, we will assess a possible role of OGT in regulating endogenous proinflammatory genes. Unlike OGT, little is known about the impact of the opposing enzyme O-Incase (OGN) on transcription. Hence, our second specific aim is to discern how these two reciprocal enzymes contribute to the activation or inhibition of the proinflammatory genetic network. Using a chromatin immunoprecipitation assay, we will study the dynamics of OGT and OGN at the promoters of either active or repressive proinflammatory genes. Moreover, we will seek to identify upstream regulators and downstream targets of these two enzymes in the transcriptional apparatus through biochemical approaches. This study is designed to delineate a precise mechanism by which O-GIcNAc regulates transcription. As glucocorticoids are currently used to treat inflammation and cancer, understanding their action via O-GIcNAc offers a new approach to treatment and drug designs.