Structural mechanism of integrin-mediated TGF-b activation

摘要：TGF-β驱动导致肺和气道纤维化的纤维炎症过程。的 本项目的长期目标是深入了解TGF-β活性的调节， 纤维化肺病的新策略和治疗方法。很少有有效的疗法来治疗慢性 肺纤维化和炎性疾病。细胞因子TGF-β是纤维化的中心介质， 病理性炎症，并且是纤维化肺病的潜在治疗靶点。然而，实际 靶向TGF-β本身或其受体的效用受到啮齿动物、灵长类动物和哺乳动物中观察到的毒性风险的限制。 人类靶向TGF-β的纤维炎性作用的更具体的方法是高度期望的。一 一种更有前景的更特异性靶向TGF-β局部作用的方法是靶向其“激活”，因为它总是 以潜伏形式（L-TGF-β）产生，必须被激活才能发挥作用。L-TGF-β的另一个特征是， 可以促进更特异性靶向的是它通过GARP共价结合到特异性细胞表面。L-TGF-β 与整联蛋白α v β 8的结合是体内TGF-β活化所必需的。对于α v β 8激活机制，如 和其他所有的研究一样，长期以来人们一直认为TGF-β必须从β细胞中释放出来，这样游离的TGF-β才能 扩散并结合靶细胞上的受体。基于最近的结构数据， 我们最近提出了一种新的模型，其中α v β 8可以结合到呈递L-TGF-β/GARP复合物的细胞上的L-TGF-β，并诱导信号传导，而不释放和扩散L-TGF-β。 TGF-β在这里，在三个目标，我们解决三个关键问题，关于这个新的模型L-TGF-β activation. (1)α v β 8/L-TGF-β/GARP三元复合物的L-TGF-β的哪个柔性结构域屏蔽TGF-β 从它的受体？(2)通过与α v β 8结合诱导的L-TGF-β的柔性是介导TGF-β所必需的吗 激活？(3)TGF-β受体（TGF-β R）是否与α v β 8/L-TGF-β/GARP复合物中的TGF-β结合？到 为了回答这些问题，我们将使用cryo-EM来确定α v β 8/L-TGF-β/GARP复合物的结构 以确定L-TGF-β的灵活性如何有助于TGF-β的激活，最后我们将捕获 TGF-β R与α v β 8/TGF-β/GARP的多聚体复合物。这些研究将提高机械 了解TGF-β激活和抑制它的治疗靶向策略。

Summary/Abstract: TGF-Beta 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-Beta 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-Beta 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-Beta 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-Beta are highly desirable. A promising method to more specifically target local effects of TGF-Beta is to target its “activation” since it is always produced in a latent form (L-TGF-Beta) that must be activated in order to function. Another feature of L-TGF-Beta that could facilitate more specific targeting is that it is covalently bound to specific cell surfaces by GARP. L-TGF-Beta binding to the integrin alphavBeta8 is essential for TGF-Beta activation in vivo. For the alphavBeta8 activation mechanism, as well as all others, it has long been assumed that TGF-Beta must be released from LAP so that free TGF-Beta 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 alphavBeta8 can bind to L-TGF-Beta on cells presenting the L-TGF-Beta/GARP complex and induce signaling without release and diffusion of TGF-Beta. Here in three aims, we address three critical questions concerning this new model of L-TGF-Beta activation. (1) Which flexible domains of L-TGF-Beta of the alphavBeta8/L-TGF-Beta/GARP ternary complex shield TGF-Beta from its receptors? (2) Is flexibility of L-TGF-Beta induced by binding to alphavBeta8 necessary to mediate TGF-Beta activation? (3) Do TGF-Beta receptors (TGF-BetaRs) bind to TGF-Beta within the alphavBeta8/L-TGF-Beta/GARP complex? To answer these questions, we will use cryo-EM to determine the structure of the alphavBeta8/L-TGF-Beta/GARP complex to determine how flexibility of L-TGF-Beta contributes to TGF-Beta activation, and finally we will capture the multimeric complex of TGF-BetaRs with alphavBeta8/LTGF-Beta/GARP. These studies will improve mechanistic understanding of TGF-Beta activation and therapeutic targeting strategies to inhibit it.