Investigation of Gene Regulation by NF-kappaB Transcription factors

描述(申请人提供)：核因子-kappaB(核因子-？B)家族的二聚体转录因子调节不同的生理过程，包括免疫和炎症反应，细胞增殖和生存。组合的核因子-β二聚体由5个家族成员组成：p50、p52、relA(P65)、cRel和RelB。这些二聚体通过与靶基因的特定DNA序列结合来调节转录，这些DNA序列称为？B位点。我们实验室的长期目标是了解核因子-B二聚体在基因调控中的具体作用。为了实现这一目标，我们已经寻求了从生物物理到遗传学的各种方法。最近的研究强调了核因子-B二聚体的翻译后修饰的作用，以及二聚体特异性共激活物在调节特定靶基因中的作用。这项建议的重点是研究通过磷酸化和共激活因子识别事件进行的转录调控。我们使用遗传学、基因组学、生物化学和生物物理学的方法来研究在人类疾病中扮演最重要角色的两个核因子-？B家族成员：RELA和P52。在目标1中，我们设计了实验来研究RelA：CBP/p300复合体的转录调控机制。一些报道已经证明了relA的磷酸化和CBP/p300结合在靶基因调控中的重要性。其他研究表明，这种修饰/相互作用的影响不是全球性的。每个修饰/相互作用只以刺激和细胞类型特定的方式影响基因的子集的表达。在本研究中，我们将研究RELA的磷酸化作用和CBP/p300的相互作用机制及其对基因调控的影响。在AIM 2中，设计了生物物理和体内实验来研究与癌症相关的p52：bcl3复合体的转录调控。作为同源二聚体，核因子-B p52和p50与bcl3结合激活转录，在免疫器官发生和耐受中发挥重要作用。其目的是研究p52同源二聚体如何与Bcl3结合的分子基础和相互作用的特异性，以及这些复合体是否靶向？B序列的一个子集。与公共卫生相关：核因子-？B二聚体转录激活剂家族，调节大量基因，对维持正常的细胞生理至关重要。这一建议的重点是了解核因子-β二聚体对启动子和转录辅助激活子识别的调节。

DESCRIPTION (provided by applicant): The NF-kappaB (NF-?B) family of dimeric transcription factors regulates diverse physiological processes including immune and inflammatory responses, cell proliferation and survival. The combinatorial NF-?B dimers are formed from five family members; p50, p52, RelA (p65), cRel and RelB. These dimers regulate transcription by binding to specific DNA sequences of target genes known as ?B sites. The long term goals of our laboratories are to understand how NF-?B dimers function specifically in gene regulation. To achieve that goal, we have pursued a variety of approaches ranging from the biophysical to the genetic. Recent studies have emphasized the role of post-translational modifications of NF-?B dimers and the role of dimer-specific co-activators in regulating specific target genes. The focus of this proposal is to examine transcriptional regulation by phosphorylation and coactivator recognition events. We employ genetic, genomic, biochemical and biophysical approaches to investigate the two NF-?B family members that play the most prominent roles in human disease: RelA and p52. In aim 1 experiments are designed to investigate the mechanisms of transcriptional control by the RelA:CBP/p300 complex. Several reports have demonstrated the importance of RelA phosphorylation, and CBP/p300 binding in target gene regulation. Other studies have revealed that the effect of this modification/interaction is not global. Each modification/interaction affects the expression of only a subset of genes in stimulus and cell type specific manner. In this study we will investigate the role of RelA phosphorylation and CBP/p300 interaction mechanism and its impact on gene regulation. In aim 2 Biophysical and in vivo experiments are designed to study the transcriptional control by the cancer-associated p52:Bcl3 complex. As homodimers, NF-?B p52 and p50 have been reported to associate with Bcl3 to activate transcription, and as such they play critical roles in immune organogenesis and tolerance. The goal of this aim is to investigate the molecular basis of how p52 homodimer associates with Bcl3 and the specificity of the interaction, and if these complexes target a subset of ?B sequences. Public Health Relevance: The NF-?B family of dimeric transcription activators, which regulates a large number of genes, is critical for maintaining normal cell physiology. The focus of this proposal is to understand the regulation of promoter and transcriptional coactivator recognition by the NF-?B dimers.