Endocytosis of Influenza Viruses

描述：（申请人提供）：病毒进入 细胞对于感染和随后的发病机制是至关重要的。对于-流感 病毒，病毒控制下的进入途径的许多方面都很好， 但病毒如何与细胞运输机制相互作用 内化和内吞作用的机制尚不清楚。已知病毒能结合 与含唾液酸的受体结合， 隔室，而是病毒在这两个事件之间如何穿过细胞，以及 哪些细胞控制机制调节这些事件是未知的。 通过细胞内吞途径的转运受GTP酶调节，例如 发动蛋白和特异性Rab蛋白。特异性抗病毒药物对病毒侵入的影响 本提案将研究这些分子的抑制作用。实验 将通过光学显微镜和超微结构水平进行， 电镜使用以下药物可抑制GTdR功能： 显性失活突变体和微量注射抑制性抗体。 我们已经证明，流感病毒进入细胞是由一种特异性的 细胞蛋白激酶C（PKC）抑制剂。这可能代表一个关键的控制 病毒进入宿主细胞的途径。的机制 该抑制剂的作用和PKC在病毒内吞中的作用将是 检查，与细胞的各种内吞途径有关。 已知流感病毒在自然界中既以大丝状形式存在， (100 nm至2 μ m）和小球形（直径100 nm）。是 被认为对人类感染很重要的丝状形式 肺，但这种形态形式的进入机制的研究， 病毒从未被传播过。然而，组织培养适应的进入 球形病毒通过网格蛋白包被的小凹发生，这似乎是一种 大型丝状病毒进入的低效途径。丝状和 球形病毒将被分离并比较它们的进入途径， 确定临床上重要的丝状病毒的进入机制。 流感仍然是人类健康的主要威胁。这些研究将导致 更全面地了解病毒的进入机制，并可能导致 开发更有效的抗病毒药物。

DESCRIPTION: (provided by applicant): The entry mechanisms of viruses into cells are critical for infection and subsequent pathogenesis. For -influenza virus, many of the aspects of the entry pathway under viral control are well understood, but how the virus interacts with the cellular trafficking machinery for internalization and endocytosis remains unclear. The virus is known to bind to a sialic acid-containing receptor and to fuse out of a low-pH intracellular compartment, but how the virus traverses the cell between these two events, and which cellular control mechanisms regulate these events are unknown. Transit through the cellular endocytic pathway is regulated by GTPases such as dynamin and specific Rab proteins. The effects on virus entry of specific inhibition of these molecules will be studied in this proposal. Experiments will be performed both by light microscopy and at the ultra-structural level by electron microscopy. GTPase function will be inhibited by use of dominant-negative mutants and by microinjection of inhibitory antibodies. We have shown that influenza virus entry into cells is inhibited by a specific inhibitor of cellular protein kinase C (PKC). This may represent a key control point for the virus in its route of entry into a host cell. The mechansim of action of the inhibitor and the role of PKC in virus endocytosis will be examined, in relation to the various endocytic pathways of the cell. Influenza virus is known to occur in nature as both a large filamentous form (100 nm by up to 2pm) and a small spherical form (100 nrn diameter). It is the filamentous forms that are thought to be important for infection in the human lung, but a study of the entry mechanism for this morphological form of the virus has never been carried out. Whereas entry of tissue culture-adapted spherical virus occurs via clathrin-coated pits, this would appear to be an inefficient route of entry for large filamentous viruses. Both filamentous and spherical viruses will be isolated and compared for their route of entry, to determine the entry mechanism for the clinically-important filamentous viruses. Influenza remains a major threat to human health. These studies will lead to a more complete understanding of the entry mechanism of the virus and may lead to the development of more effective anti-viral drugs.