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Protein interactions regulating HIV replication in macrophages

调节巨噬细胞中 HIV 复制的蛋白质相互作用

基本信息

批准号：
10667478
负责人：
George Kyei
金额：
$ 39.38万
依托单位：
WASHINGTON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-08-03 至 2025-07-31
项目状态：
未结题

项目摘要

ABSTRACT Macrophages serve as HIV reservoir, which is a barrier to cure. However, the molecular mechanism underlying how the virus establishes and maintains infection in this cell type is much less understood. Elucidating the mechanisms of HIV replication in macrophages is of vital importance to the HIV eradication efforts. We previously demonstrated that cyclin L2 is required for HIV-1 infection in macrophages, interacts with and targets the HIV restriction factor SAM domain and HD domain-containing protein 1 (SAMHD1) for degradation. However, the details of the cyclin L2-SAMHD1 interactions are unknown. In addition, how cyclin L2 is regulated in macrophages is unknown. In our recent studies we showed that cyclin L2 interacts with and is phosphorylated by the Dual Specificity Tyrosine Phosphorylation Regulated Kinase 1A (DYRK1A). While knockout of cyclin L2 decreased HIV infection 10-fold, depletion of DYRK1A increased HIV infection in primary macrophages up to 10-fold. The overarching objective of this proposal is to determine how the interplay between cyclin L2, DYRK1A and SAMHD1 regulate HIV replication in macrophages. Our specific aims are: Aim 1: Define the mechanism by which the kinase DYRK1A regulates cyclin L2 in macrophages. We hypothesize that DYRK1A inhibits cyclin L2 through phosphorylation or degradation. First we will determine whether DYRK1A-mediated phosphorylation of cyclin L2 prevents the promotion of HIV infection in macrophages. Second, we will define the molecular domains of cyclin L2 and DYRK1A required for their interactions. Third, we will determine the mechanism by which DYRK1A regulate cyclin L2 levels in macrophages. Aim 2: Determine how cyclin L2 and HIV-1 regulate SAMHD1 in macrophages during viral infection. HIV- 1 possesses no Vpx, but is able to establish infection in macrophages despite the abundance of SAMHD1. Therefore, HIV may orchestrate the degradation of SAMHD1 through cyclin L2 or other means. In this aim, we will (i) determine the molecular domains of cyclin L2 and SAMHD1 required for this interaction (ii) identify the viral or cellular factor(s) that induce elevation and reduction in cyclin L2 and SAMHD1 levels respectively during early HIV infection and (iii) determine if SAMHD1 degradation in macrophages is influenced by cyclin L2 phosphorylation by DYRK1A. These studies will reveal how the cyclin L2-DYRK1A-SAMHD1 complex regulate HIV infection in macrophages. Our efforts will be highly significant for understanding a novel pathway that determines whether HIV replication is restricted or enabled in macrophages. The potential impact is significant since the dissection of these protein interactions could reveal potential therapeutic targets against HIV infection.

摘要巨噬细胞是HIV的储存库，是治愈的障碍。然而，潜在的分子机制病毒如何在这种细胞类型中建立和维持感染还不太清楚。阐明 HIV在巨噬细胞中复制的机制对于HIV根除工作至关重要。我们以前证明，细胞周期蛋白L2是HIV-1感染巨噬细胞所必需的，靶向HIV限制因子SAM结构域和含HD结构域的蛋白1（SAMHD 1）进行降解。然而，细胞周期蛋白L2-SAMHD 1相互作用的细节是未知的。此外，细胞周期蛋白L2是如何被调节的，在巨噬细胞中是未知的。在我们最近的研究中，我们发现细胞周期蛋白L2与细胞周期蛋白L2相互作用并被磷酸化，双特异性酪氨酸磷酸化调节激酶1A（DYRK 1A）。敲除细胞周期蛋白L2 减少HIV感染10倍，DYRK 1A的耗竭增加了原代巨噬细胞中的HIV感染，十倍本提案的首要目标是确定细胞周期蛋白L2、DYRK 1A 和SAMHD 1调节HIV在巨噬细胞中的复制。我们的具体目标是：目的1：阐明DYRK 1A激酶对巨噬细胞周期蛋白L2的调控机制。我们推测DYRK 1A通过磷酸化或降解抑制细胞周期蛋白L2。首先我们要确定 DYRK 1A介导的细胞周期蛋白L2磷酸化是否阻止了HIV感染的促进，巨噬细胞其次，我们将定义细胞周期蛋白L2和DYRK 1A的分子结构域，交互.第三，我们将确定DYRK 1A调节细胞周期蛋白L2水平的机制。巨噬细胞目的2：确定细胞周期蛋白L2和HIV-1在病毒感染过程中如何调节巨噬细胞中的SAMHD 1。艾滋病毒- 1不具有Vpx，但能够在巨噬细胞中建立感染，尽管SAMHD 1丰富。因此，HIV可能通过细胞周期蛋白L2或其他方式协调SAMHD 1的降解。为此，我们将（i）确定这种相互作用所需的细胞周期蛋白L2和SAMHD 1的分子结构域（ii）鉴定病毒或细胞因子，其分别诱导细胞周期蛋白L2和SAMHD 1水平的升高和降低，早期HIV感染和（iii）确定巨噬细胞中SAMHD 1降解是否受细胞周期蛋白L2影响 DYRK 1A的磷酸化。这些研究将揭示细胞周期蛋白L2-DYRK 1A-SAMHD 1复合物如何调节巨噬细胞中的HIV感染。我们的努力对于理解一种决定艾滋病毒复制是否在巨噬细胞中被限制或激活。潜在的影响是显着的，因为这些蛋白质的解剖相互作用可能揭示潜在的治疗艾滋病毒感染的目标。