Molecular Basis for Regulation of Cellular Stress Response Pathways by CBP/p300

转录共激活物CREB结合蛋白(CBP)和p300调节多种信号 真核细胞中激活或抑制基因表达的转导途径。他们是 对于诸如生长、分化、凋亡、DNA修复和 胚胎发育。它们还发挥着肿瘤抑制因子的作用，并调节控制 细胞增殖、肿瘤发生和癌症进展。CBP和p300在 许多信号通路调节对细胞应激的反应，从而在细胞内发挥关键作用 命运决定-决定细胞是应该存活和增殖，还是应该经历凋亡。CBP和 P300是细胞信号电路中的中心中枢；细胞中CBP/P300的数量是有限的 而调节蛋白必须竞争结合。CBP和p300是激活所必需的 肿瘤抑制基因P53和P53介导的应激反应通路的调节。他们也 通过激活氧应激的转录，在对缺氧的反应中发挥不可或缺的作用 受缺氧诱导转录因子HIF-1α控制的基因，以及作为中枢的基因 CITED2蛋白下调缺氧诱导因子-1α反式激活的负反馈回路 CBP/P300的TAZ1结构域的竞争。本提案的首要目标是 阐明CBP和p300发挥中枢调节作用的结构和分子基础 控制低氧反应，防止致癌转化。初步数据显示 CBP/p300TAZ1结构域和HIF-1α和CITED2的激活结构域具有功能 合作创造了一种单向超敏分子开关，有效地取代了HIF- 1CBP/p300中的α激活结构域下调HIF-1α反应基因的转录。这个 驱动这种转换的分子间相互作用将通过亲和力测量和 停流动力学，以及系统的结构和动态特征将被表征为 核磁共振提供了对这种超敏变构的详细分子机制的见解 交换机功能。在p65亚基之间的竞争中也观察到了类似开关的行为 核转录因子κB和缺氧诱导因子-1α，P53和P65之间的结合。生物物理测量将是 确定p65、P53和HIF-1α竞争机制。鉴于的核心作用 CBP/p300作为关键细胞信号网络中浓度受限的中枢，很可能是变构 可能是一种无序的信号蛋白竞争靶标的一般机制。

The transcriptional coactivators CREB binding protein (CBP) and p300 regulate numerous signal transduction pathways in eukaryotic cells by activation or repression of gene expression. They are essential for such basic cellular functions as growth, differentiation, apoptosis, DNA repair, and embryonic development. They also function as tumor suppressors and regulate key genes that control cellular proliferation, tumorigenesis, and cancer progression. CBP and p300 mediate crosstalk between many signaling pathways that regulate the response to cellular stress, thereby playing a key role in cell fate decisions – determining whether cells should survive and proliferate or undergo apoptosis. CBP and p300 are central hubs in the signaling circuitry of the cell; the amount of CBP/p300 in the cell is limiting and regulatory proteins must compete for binding. CBP and p300 are absolutely required for activation of the tumor suppressor p53 and regulation of p53-mediated stress response pathways. They also perform an indispensable function in the response to hypoxia, by activating transcription of oxygen stress genes controlled by the hypoxia inducible transcription factor HIF-1α and by acting as a central hub in a negative feedback circuit in which the protein CITED2 downregulates HIF-1α transactivation by competition for the TAZ1 domain of CBP/p300. The overarching goals of the present proposal are to elucidate the structural and molecular basis by which CBP and p300 perform their central regulatory roles to control the hypoxic response and to protect against oncogenic transformation. Preliminary data show that the CBP/p300 TAZ1 domain and the activation domains of HIF-1α and CITED2 function cooperatively to create a unidirectional hypersensitive molecular switch that efficiently displaces the HIF- 1α activation domain from CBP/p300 to downregulate transcription of HIF-1α responsive genes. The intermolecular interactions that drive this switch will be investigated using affinity measurements and stopped flow kinetics, and the structural and dynamic features of the system will be characterized by NMR to provide insights into the detailed molecular mechanism by which this hypersensitive allosteric switch functions. Switch-like behavior has also been observed in competition between the p65 subunit of NFκB and HIF-1α, and between p53 and p65 for binding to TAZ1. Biophysical measurements will be performed to determine the mechanism of p65, p53, and HIF-1α competition. Given the central role of CBP/p300 as concentration-limited hubs in critical cellular signaling networks, it is likely that allostery may be a general mechanism by which disordered signaling proteins compete for their targets.