Virological Synapse and Signaling for Efficient HIV Transmission

描述(由申请人提供)： 与许多其他病毒(如HTLV-1和疱疹病毒)一样，艾滋病毒通过细胞间传播比无细胞病毒传播更有效地从感染细胞传播到新的靶细胞。细胞间的传播发生在感染性或病毒学突触(VS)中，感染细胞和靶细胞之间的紧密裂隙是由于两个相对细胞的质膜牢固附着的结果。VS的形成始于HIV包膜gp120与CD4的相互作用，然后招募特定的细胞膜蛋白，包括相关的辅助受体(CCR5或CXCR4)和细胞黏附分子。重要的是，VS还需要细胞内信号分子在表达gp120的供体细胞和CD4靶细胞中的积极参与。我们最近的研究表明，在靶T细胞(“传出”)侧，一系列信号事件被触发，从激活CD4相关的Lck开始，导致T细胞运动能力的改变和肌动蛋白细胞骨架的重组，其中每一项都可能对VS的HIV转移和感染的早期阶段的成功进展起重要作用。在受感染的供体细胞(“传入”)一侧，Lck也促进了病毒的组装和萌发，它将HIV Gag结合并引导到受感染T细胞的质膜上。信号分子在VS形成中的重要性为利用这些信号转导途径阻断HIV传播和感染提供了机会。在这项应用中，我们将关注HIV从感染的CD4T细胞向靶CD4T细胞的传播，这是HIV感染的最常见的细胞类型，并定义在感染的供体细胞和未感染的靶细胞中激活的膜近端信号分子和通路，这些分子和通路是VS形成所需的。识别这些分子和途径将使我们有机会抑制它们的活性，阻止艾滋病毒通过VS传播。为了实现这一目标，我们将评估LCK激活及其下游信号事件对HIV有效的VS形成和细胞间传播的重要性(目标1)。我们将确定在感染的供体细胞中招募到VS的膜-近端信号分子，并将它们与在靶T细胞中发现的信号分子进行比较(目标2)。还将研究Lck和其他激活信号在细胞骨架重组中的作用，以促进VS介导的病毒外泄和进入(目标3)。最后，我们将确定HIV Env与靶CD4T细胞上的1427整合素相互作用如何通过VS以及VS形成和信号传递影响HIV传播(目标4)。VS形成所必需的信号分子和信号通路的鉴定将为信号转导抑制剂作为一类新的HIV抑制剂的发展铺平道路。人们对这种抗艾滋病毒模式越来越感兴趣，因为它们绕过了艾滋病毒-1基因和抗原变异这一前所未有的问题，该问题阻碍了有效艾滋病毒/艾滋病疫苗的开发，并导致了抗药性病毒的出现。

DESCRIPTION (provided by applicant): Summary HIV, like many other viruses (e.g. HTLV-1 and herpes viruses), disseminates from infected cells to new target cells much more efficiently by cell-to-cell transmission as compared to cell-free virion transmission. Cell- to-cell transmission occurs in an infectious or virological synapse (VS), where a tight cleft between an infected cell and a target cell is formed as a result of firmly adhering plasma membranes of the two apposing cells. VS formation is initiated upon HIV envelope gp120 interaction with CD4 which then recruits specific cell membrane proteins including the relevant co-receptor (CCR5 or CXCR4) and cellular adhesion molecules. Importantly, VS also requires active participation of intracellular signaling molecules in both the gp120-expressing donor cell and the CD4+ target cell. Our recent studies demonstrate that in the target T cell ("efferent") side, a cascade of signaling events are triggered, starting from activation of CD4-associated Lck, to cause alteration of T cell motility and reorganization of actin cytoskeleton, each of which is potentially important for successful progression of the early stages of HIV transfer and infection at the VS. In the infected donor cell ("afferent") side, virus assembly and budding are also facilitated by Lck, which binds and directs HIV gag to the plasma membrane in the infected T cell. The importance of signaling molecules in VS formation presents an opportunity to exploit these signal transduction pathways for blocking HIV transmission and infection. In this application we will focus on HIV transmission from infected CD4 T cells to target CD4 T cells, the most common cell type infected by HIV, and define the membrane-proximal signaling molecules and pathways activated in the infected donor and uninfected target cells that are required for VS formation. Identifying these molecules and pathways will allow us the opportunity to inhibit their activity to stop HIV transmission through the VS. To achieve this, we will evaluate the importance of Lck activation and its downstream signaling events for efficient VS formation and cell-cell transmission of HIV (Aim 1). We will determine membrane-proximal signaling molecules that are recruited to VS in the infected donor cells and compare them with those found in the target T cells (Aim 2). The involvement of Lck and other activation signals in cytoskeleton reorganization that facilitates VS-mediated virus egress and entry will also be studied (Aim 3). Finally, we will determine how HIV Env interaction with the 1427 integrin on the target CD4 T cells affects HIV transmission via the VS, and VS formation and signaling (Aim 4). Identification of signaling molecules and pathways essential for VS formation should pave the way for the development of signal transduction inhibitors as a novel class of HIV inhibitors. There is a growing interest for such anti-HIV modalities as they circumvent the unprecedented problem of HIV-1 genetic and antigenic variability, which has hampered the development of efficacious HIV/AIDS vaccines and has led to the emergence of drug-resistant viruses.