Molecular basis of selectivity for coactivator recruitment by ARNT PAS domains

描述（由申请人提供）：异常细胞生长产生缺氧微环境，用于抑制增殖;肿瘤通过缺氧反应途径的组成性激活来规避这种稳态调节，缺氧反应途径上调厌氧代谢和血管生成基因，以促进细胞生长，否则细胞生长将受到低氧的限制。缺氧反应途径由氧敏感性HIF-α（缺氧诱导因子-α）亚基和ARNT（芳烃核转运蛋白）进行转录调控，它们形成异二聚体bHLH-PAS转录激活因子，结合DNA中的靶序列并招募共激活因子启动转录。本研究的目的是通过研究ARNT PAS结构域募集辅激活因子的结构和功能基础，探讨ARNT在转录激活中的作用。我们假设ARNT PAS结构域通过利用这个小的模块化结构域上的相对界面同时介导与HIF-a和共激活因子的相互作用。在目标1中，我们将确定ARNT：辅激活因子相互作用的最小基序，并定义ARNT的辅激活因子特异性的分子基础。在目标2中，我们将确定ARNT的结构基础：共激活剂相互作用和功能的复杂破坏在体内的后果。在目标3中，我们将筛选ARNT PAS结构域的人工配体，以确定破坏ARNT：辅激活因子相互作用的小分子拮抗剂。这项工作将研究ARNT如何有助于转录调节，以应对缺氧，并确定化合物，阻止ARNT功能的潜在治疗价值，在防止肿瘤生长。

DESCRIPTION (provided by applicant): Aberrant cellular growth creates oxygen-poor microenvironments that serve to inhibit proliferation; tumors circumvent this homeostatic regulation through constitutive activation of the hypoxia response pathway, which up-regulates genes for anaerobic metabolism and angiogenesis to facilitate cellular growth that would otherwise be limited by low oxygen. The hypoxia response pathway is transcriptionally regulated by an oxygen-sensitive HIF-a (hypoxia-inducible factor-a) subunit and ARNT (aryl hydrocarbon nuclear translocator), which form a heterodimeric bHLH-PAS transcriptional activator that binds target sequences in DNA and recruits coactivators to initiate transcription. The objective of this proposal is to investigate the role of ARNT in transcriptional activation by examining the structural and functional basis of coactivator recruitment by ARNT PAS domains. We hypothesize that ARNT PAS domains simultaneously mediate interaction with HIF-a and coactivators by utilizing opposing interfaces on this small, modular domain. In Aim 1, we will identify minimal motifs for ARNT:coactivator interactions and define the molecular basis of coactivator specificity for ARNT. In Aim 2, we will determine the structural basis for the ARNT:coactivator interaction and characterize functional consequences of complex disruption in vivo. In Aim 3, we will screen ARNT PAS domains for artificial ligands to identify small molecule antagonists that disrupt the ARNT:coactivator interaction. This work will examine how ARNT contributes to transcriptional regulation in response to hypoxia and identify compounds that block ARNT function with potential therapeutic value in preventing tumor growth.