Structural mechanism of integrin-mediated TGF-b activation

摘要/摘要：转化生长因子-驱动纤维炎症过程，导致肺和呼吸道纤维化。这个 本项目的远期目标是深入了解转化生长因子-活性的调控发展 纤维化肺病的新策略和新疗法。治疗慢性阻塞性肺疾病的有效方法很少 肺纤维化和炎症性疾病。细胞因子转化生长因子-是肝纤维化和 是一种病理性炎症，是治疗纤维化肺疾病的潜在靶点。然而，实际情况是 靶向转化生长因子-本身或其受体的效用受到啮齿动物、灵长类动物和 人类。针对转化生长因子-的纤维炎症效应的更具体的方法是非常可取的。一个 靶向转化生长因子-的局部效应的有希望的方法是靶向其“激活”，因为它总是 以潜伏形式产生(L-转化生长因子-)，必须激活才能发挥作用。L-转化生长因子-的另一个特点是 可以促进更具体的靶向是它通过GARP共价结合到特定的细胞表面。L-转化生长因子- 与整合素v8的结合是体内转化生长因子激活所必需的。对于v8激活机制，AS 和所有其他的一样，长期以来人们一直认为转化生长因子-必须从LAP中释放出来，这样游离的转化生长因子-才能 将其受体扩散并结合到靶细胞上。基于使用单粒子获得的最新结构数据 电子冷冻显微镜(Cryo-EM)，我们最近提出了一种新的v8与L结合的模型- 转化生长因子-在呈现L-转化生长因子-/GARP复合体的细胞上诱导信号转导而不释放和扩散 转化生长因子-。在这里，我们以三个目标来解决关于L-转化生长因子-这一新模型的三个关键问题 激活。(1)L-转化生长因子-的v8/L-转化生长因子-/GARP三元络合物的哪些柔性结构域屏蔽转化生长因子- L-转化生长因子-与--8结合诱导的柔韧性是介导转化生长因子-所必需的吗？ (3)转化生长因子-受体(-R)是否与转化生长因子-在v8/L-转化生长因子-/GARP复合体中结合？至 回答这些问题，我们将使用冷冻EM来确定v8/L-转化生长因子-/GARP复合体的结构 为了确定L-转化生长因子-的灵活性如何促进转化生长因子-的激活，最后我们将捕获 转化生长因子-受体与v8//GARP的多聚体复合物这些研究将改进机械学 了解转化生长因子-的激活和抑制它的治疗靶向策略。

Summary/Abstract: TGF- drives the fibroinflammatory processes that leads to lung and airway fibrosis. The long-term goal of this project is to acquire a deep understanding of the regulation of TGF- activity to develop new strategies and treatments for fibrosing lung disease. There are few effective therapies to treat chronic fibrosing and inflammatory diseases of the lung. The cytokine TGF- is a central mediator of fibrosis and pathologic inflammation and is a potential therapeutic target in fibrosing lung disease. However, the practical utility of targeting TGF- itself or its receptors is limited by risk of toxicities seen in rodents, primates and humans. More specific methods to target the fibroinflammatory effects of TGF- are highly desirable. A promising method to more specifically target local effects of TGF- is to target its “activation” since it is always produced in a latent form (L-TGF-) that must be activated in order to function. Another feature of L-TGF- that could facilitate more specific targeting is that it is covalently bound to specific cell surfaces by GARP. L-TGF- binding to the integrin v8 is essential for TGF- activation in vivo. For the v8 activation mechanism, as well as all others, it has long been assumed that TGF- must be released from LAP so that free TGF- can diffuse and bind its receptors on target cells. Based on recent structural data obtained using single particle electron cryomicroscopy (cryo-EM), we have recently proposed a new model whereby v8 can bind to L- TGF- on cells presenting the L-TGF-/GARP complex and induce signaling without release and diffusion of TGF-. Here in three aims, we address three critical questions concerning this new model of L-TGF- activation. (1) Which flexible domains of L-TGF- of the v8/L-TGF-/GARP ternary complex shield TGF- from its receptors? (2) Is flexibility of L-TGF- induced by binding to v8 necessary to mediate TGF- activation? (3) Do TGF- receptors (TGF-Rs) bind to TGF- within the v8/L-TGF-/GARP complex? To answer these questions, we will use cryo-EM to determine the structure of the v8/L-TGF-/GARP complex to determine how flexibility of L-TGF- contributes to TGF- activation, and finally we will capture the multimeric complex of TGF-Rs with v8/LTGF-/GARP. These studies will improve mechanistic understanding of TGF- activation and therapeutic targeting strategies to inhibit it.