HIV Release and Restriction

摘要 本项目旨在阐明HIV-1出芽及其被宿主抑制的机制基础 因子retroCHMP 3.为了启动出芽，HIV-1 Gag结合ESCRT-I1-4和ALIX 5 -7，这两个早期作用因子在HIV-1的细胞中起作用。 转运所需的宿主内体分选复合物（ESCRT）途径。这些因素，反过来，招募 晚期作用的ESCRT因子CHMP 2、CHMP 4（ESCRT-III）和VPS 4（AAA ATP酶）2，8 -10（在参考文献中综述。 11-16）。已经提出了许多膜收缩和分裂的模型。17 -26我们倾向于那些在 其中：1）ESCRT-III蛋白形成螺旋丝，招募VPS 4并收缩膜颈， 2）VPS 4 ATP酶通过重塑潜在的ESCRT-III促进脂质混合 脚手架我们现在建议确定ESCRT-III丝膜重塑的结构基础 和VPS 4（目的1和2），表征哺乳动物retroCHMP 3（ESCRT-III）蛋白如何抑制HIV 出芽而不诱导细胞毒性（目标3），并通过以下方法对出芽病毒粒子内的ESCRT组装体进行成像： 电子冷冻断层扫描（Aim 4）。这些研究将揭示HIV-1如何利用宿主ESCRT-III/VPS 4 以及宿主retroCHMP 3蛋白如何特异性抑制该过程。

ABSTRACT This Project is designed to elucidate the mechanistic bases for HIV-1 budding and its inhibition by the host factor retroCHMP3. To initiate budding, HIV-1 Gag binds ESCRT-I1-4 and ALIX5-7, two early-acting factors in the host Endosomal Sorting Complexes Required for Transport (ESCRT) pathway. These factors, in turn, recruit the late-acting ESCRT factors CHMP2, CHMP4 (ESCRT-III) and VPS4 (AAA ATPase)2,8-10 (reviewed in refs. 11-16). Numerous models for membrane constriction and fission have been proposed.17-26 We favor those in which: 1) ESCRT-III proteins form spiraling filaments that recruit VPS4 and constrict the membrane neck from the cytoplasmic face and 2) The VPS4 ATPase promotes lipid mixing by remodeling the underlying ESCRT-III scaffold. We now propose to determine the structural bases for membrane remodeling by ESCRT-III filaments and VPS4 (Aims 1 and 2), characterize how mammalian retroCHMP3 (ESCRT-III) proteins can inhibit HIV budding without inducing cellular toxicity (Aim 3), and image ESCRT assemblies within budding virions by electron cryotomography (Aim 4). These studies will reveal how HIV-1 uses the host ESCRT-III/VPS4 machinery to exit cells and how host retroCHMP3 proteins can specifically inhibit this process.