Nef-Host Interactions in Virus Replication

描述(由申请人提供)：NEF是一种辅助病毒基因产物，已被证明是猴子模型中高滴度病毒复制和艾滋病发病所必需的。此外，感染nef缺陷的HIV-1毒株与人类没有疾病进展有关。虽然Nef似乎对体内有效的病毒复制至关重要，但它的作用机制却鲜为人知。在细胞培养中，Nef可以下调细胞表面的CD4和MHC-I类分子，调节细胞的激活状态，并直接增强后代病毒粒子的感染力。我们的初步结果表明，HIV-1Nef以一种高度特异的方式与普遍表达的GTP酶Dynamin 2相互作用，Dynamin 2对于网状蛋白介导的内吞作用是必不可少的，并在胞内囊泡萌发过程中介导分裂步骤。我们的结果还表明，这种相互作用对于Nef增强病毒感染力是至关重要的，因为我们发现这一功能可以被显性负Dynamin 2特异性抑制，但不能被Dynamin 1特异性抑制。由于Dynamin 2参与内吞作用，我们的结果进一步表明，Nef可能需要下调细胞表面一个尚未确定的因子，以获得最佳的HIV-1感染力。我们的目标是阐明是什么使Nef能够与Dynamin 2相互作用，而不是与密切相关的异构体相互作用，了解这种相互作用与HIV-1感染性和复制的相关性，并确定Nef和Dynamin 2的下游靶点。了解Dynamin 2在增强Nef感染性中的作用最终可能为抗病毒治疗提供有希望的新靶点，特别是因为已知的Dynamin 2功能表明参与了细胞表面蛋白的参与。

DESCRIPTION (provided by applicant): Nef is an accessory viral gene product that has been shown to be required for high titer virus replication and AIDS pathogenesis in a monkey model. Furthermore, infection with nef-defective strains of HIV-1 has been associated with the absence of disease progression in humans. While Nef appears crucial for efficient virus replication in vivo, the mechanism by which it acts is poorly understood. In cell culture, Nef can be shown to down-regulate CD4 and MHC class I from the cell surface, to modulate the cellular activation state, and to directly enhance the infectivity of progeny virions. Our preliminary results now establish that HIV-1 Nef interacts in a highly specific manner with the ubiquitously expressed GTPase dynamin 2, which is essential for clathrin-mediated endocytosis and mediates the fission step during endocytic vesicle budding. Our results also imply that this interaction is crucial for the enhancement of viral infectivity by Nef, since we find that this function can be specifically inhibited by dominant-negative dynamin 2 but not dynamin 1. Because of the involvement of dynamin 2 in endocytosis, our results further suggest that the down-regulation of an as yet unidentified factor from the cell surface by Nef may be required for optimal HIV-1 infectivity. Our goals are to elucidate what enables Nef to interact with dynamin 2 but not with closely related isoforms, to understand the relevance of the interaction for HIV-1 infectivity and replication, and to identify the downstream target of Nef and dynamin 2. An understanding of the role of dynamin 2 in the infectivity enhancement of Nef may ultimately provide promising new targets for anti-viral therapy, particularly since the known function of dynamin 2 suggests the involvement of a cell surface protein.