Allosteric regulation of SIRT1 by a PACS-2 and DBC1 regulatory hub

项目摘要 “营养感应”酶SIRT 1位于一系列复杂的分子相互作用的十字路口， 影响肥胖、糖尿病、神经变性和癌症等多种疾病的易感性。一 治疗肥胖相关疾病的有希望的治疗策略包括小分子STAC， 其结合SIRT 1 N-末端区域以变构地增加SIRT 1活性，从而提高SIRT 1的活性。 分解代谢PGC-1 β/PPAR β靶基因的表达有助于防止饮食诱导的肥胖。STAC 在位于SIRT 1 N-末端区域的远端部分的3-螺旋束中结合SIRT 1。这个3螺旋束是 由SIRT 1 N-末端区域的上游元件屏蔽，保护酶免受未鉴定的 细胞调节器因此，通过与蛋白质相互作用来控制酶活性的细胞蛋白质的鉴定， 3-螺旋束是理解SIRT 1调节的关键。我们公布的和初步的数据表明 PACS-2是一种这样的SIRT 1调节器，PACS-2、DBC 1和SIRT 1形成了一个新的三方枢纽， 控制SIRT 1脱乙酰酶活性。DBC 1与SIRT 1的N-末端区域结合，在那里它促进PACS-2 募集和结合SIRT 1。PACS-2使关键的3-螺旋束不稳定，从而抑制SIRT 1活性 并因此抑制PGC-1 β/PPAR β靶基因的SIRT 1依赖性活化。我们的长期目标 是为了了解SIRT 1是如何被调节来控制人类的能量稳态的。的目的 具体应用是确定PACS-2和DBC 1如何协同作用以变构调节SIRT 1 酶的活性以及这个调节中心如何控制体内对禁食或营养过剩的反应。我们 中心假设是SIRT 1/PACS-2或SIRT 1/DBC 1相互作用的破坏将阻止高脂血症， 饮食诱导的SIRT 1抑制，因此保护小鼠免受肝脂肪变性和胰岛素抵抗。 在强有力的初步数据的指导下，我们将通过追求三个具体目标来测试我们的假设：1）确定 PACS-2/SIRT 1相互作用调节酶活性的构象和机制步骤; 2） 确定PACS-2和DBC 1的协同作用如何抑制SIRT 1;以及3）确定PACS-2和DBC 1的生理作用。 PACS-2、DBC 1和SIRT 1之间的相互作用对肝脏代谢的重要性。该方法是 创新，因为它将表征，从原子结构到整个有机体功能， 在禁食和营养过剩期间控制能量代谢的未被识别的调节中枢。本研究是 重要的是，它将揭示SIRT 1调节剂如何控制酶的活性，以及我们如何影响 其功能是改善健康。

PROJECT SUMMARY The “nutrient-sensing” enzyme SIRT1 lays at the crossroads of a complex array of molecular interactions that impact susceptibility to diseases as diverse as obesity, diabetes, neurodegeneration, and cancer. One promising therapeutic strategy for the treatment of obesity-related morbidity involves small-molecule STACs, which bind the SIRT1 N-terminal region to allosterically increase SIRT1 activity, thereby elevating the expression of catabolic PGC-1/PPAR target genes that help to protect against diet-induced obesity. STACs bind SIRT1 in a 3-helix bundle located at the distal part of the SIRT1 N-terminal region. This 3-helix bundle is shielded by an upstream element of the SIRT1 N-terminal region, protecting the enzyme from an unidentified cellular regulator. Thus, identification of cellular proteins that control enzyme activity through interaction with the 3-helix bundle is key to understanding SIRT1 regulation. Our published and preliminary data suggest PACS-2 is one such SIRT1 regulator and that PACS-2, DBC1 and SIRT1 form a novel tripartite hub that controls SIRT1 deacetylase activity. DBC1 binds to the N-terminal region of SIRT1, where it promotes PACS-2 recruitment and binding to SIRT1. PACS-2 destabilizes the critical 3-helix bundle, which inhibits SIRT1 activity and consequently represses SIRT1-dependent activation of PGC-1/PPAR target genes. Our long-term goal is to understand how SIRT1 is regulated to control energy homeostasis in humans. The objective of this particular application is to determine how PACS-2 and DBC1 synergize to allosterically modulate SIRT1 enzyme activity and how this regulatory hub controls the response to fasting or overnutrition in vivo. Our central hypothesis is that disruption of the SIRT1/PACS-2 or SIRT1/DBC1 interactions will prevent high-fat- diet-induced repression of SIRT1 and, therefore, protect mice from hepatic steatosis and insulin resistance. Guided by strong preliminary data, we will test our hypothesis by pursuing three specific aims: 1) Determine the conformational and mechanistic steps by which the PACS-2/SIRT1 interaction regulates enzyme activity; 2) Determine how synergistic actions by PACS-2 and DBC1 inhibit SIRT1; and 3) Determine the physiologic importance of interactions between PACS-2, DBC1, and SIRT1 on hepatic metabolism. The approach is innovative because it will characterize, from the atomic structure to whole-organism function, a previously unrecognized regulatory hub controlling energy metabolism during fasting and overnutrition. This research is significant because it will uncover how SIRT1 regulators control enzyme activity and how we can influence their functions to improve health.