Modulation of Rab7-Dependent Degradative Pathways by Novel Adenovirus Protein

内吞作用有许多重要的功能，包括获取细胞外 调节细胞表面受体表达和信号转导的营养物质， 维持细胞极性和抗原呈递。许多细胞内病原体 劫持内吞途径，以侵入细胞、增殖和确保病原体 生死存亡。对病原性感染后果的了解使我们能够 对细胞内转运机制的了解。分离的致病基因有 也被用来抑制与其细胞靶标相关的病理过程 人类疾病模型。这一提议的重点是理解分子和 早期编码的完整膜蛋白所使用的细胞机制 人类腺病毒的第3区称为RID？，最初发现是因为 其将结构性循环的EGF受体转移到溶酶体的能力。我们有 最近发现RID？与RILP和ORP1L相互作用，这两个已知的效应器 Rab7 GTP酶调控着从早期到晚期的内切体的转运，然后到 溶酶体。重要的是，RID？弥补了Rab7功能的丧失，这表明 通过协调Rab7效应器的募集来改变内吞体膜动力学 到通常被认为是早期分选内小体的隔室。致我们的 知道这是第一个由DNA病毒编码的蛋白质，通过Rab7发挥作用 模仿。与其他小的GTP酶类似，Rab7的作用方式是在活性的GTP- 绑定状态和非活动的GDP绑定状态。相比之下，RID？是一种非酶 与Rab7缺乏任何序列同源性的固有膜蛋白，提供了一种 汇聚进化的显著例子。四个具体的目标将检验这些 假设。1)RID通过招募RILP来控制微管依赖的囊泡运输 和ORP1L，然后激活负末端定向的动力蛋白-动力蛋白马达。2)。RID？- RILP促进ESCRT-II依赖的EGF受体不依赖受体的分选 酪氨酸激酶或泛素状态。3)RID？-ORP1L调节胆固醇流出 维持细胞内适当的脂类平衡所必需的内小体。4)RID？ 补偿高效性腺病毒感染期间Rab7功能的丧失，以及 通过干扰肿瘤坏死因子？-表皮生长因子来钝化腺病毒诱导的炎症性疾病 调节IL-8产生的受体信号级联反应。总之，这些研究将 为膜蛋白运输的细胞生物学提供了新的见解，而且 腺病毒引起炎症性疾病的分子基础。

Endocytosis serves many important functions ranging from acquisition of extracellular nutrients to regulation of cell surface receptor expression and signal transduction, maintenance of cell polarity, and antigen presentation. Many intracellular pathogens hijack endocytic pathways in order to invade cells, proliferate, and ensure pathogen survival. Understanding the consequences of pathogenic infection has enabled greater understanding of the endocytic trafficking machinery. Isolated pathogenic genes have also been used to curb pathological processes associated with their cellular targets in human disease models. The focus of this proposal is to understand the molecular and cellular mechanisms employed by an integral membrane protein encoded by the early region 3 of human adenoviruses called RID¿, which was originally identified because of its ability to divert constitutively recycling EGF receptors to lysosomes. We have recently discovered that RID¿ interacts with RILP and ORP1L, two known effectors for the Rab7 GTPase that governs transport from early to late endosomes and then to lysosomes. Importantly, RID¿ compensates for Rab7 loss-of-function, suggesting it modifies endosome membrane dynamics by coordinating recruitment of Rab7 effectors to compartments that would ordinarily be perceived as early sorting endosomes. To our knowledge this is the first protein encoded by a DNA virus that functions by Rab7 mimicry. Similar to other small GTPases, Rab7 acts by cycling between an active GTP- bound state and an inactive GDP-bound state. In contrast RID¿ is a non-enzymatic intrinsic membrane protein that lacks any sequence homology to Rab7, providing a remarkable example of convergent evolution. Four specific aims will test these hypotheses. 1) RID¿ controls microtubule-dependent vesicle transport by recruiting RILP and ORP1L which then activate minus end-directed dynein-dynactin motors. 2). RID¿- RILP facilitates ESCRT-II-dependent EGF receptor sorting independent of receptor tyrosine kinase or ubiquitin status. 3) RID¿-ORP1L regulates cholesterol efflux from endosomes necessary to maintain the proper lipid balance in cells. 4) RID¿ compensates for Rab7 loss-of-function during a productive adenovirus infection, and blunts adenovirus-induced inflammatory disease by interfering with a TNF¿-EGF receptor signaling cascade that regulates IL-8 production. Altogether these studies will provide novel insights to the cell biology of membrane protein trafficking, and also the molecular basis for adenovirus-induced inflammatory disease.