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

• 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
• 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
• 来源: https://reporter.nih.gov/project-details/10667478
• 关键词: C-terminal; CD4 Positive T Lymphocytes; Cells; Complex; Cyclins; Data; Dissection; Goals; HIV; HIV Infections; HIV-1; HIV-2; Hour; Infection; Knock-out; Macrophage; Mass Spectrum Analysis; Mediating; Molecular; Mutagenesis; Nucleotides; Pathway interactions; Phosphorylation; Phosphorylation Site; Phosphotransferases; Production; Proteins; Rest; SAM Domain; Specificity; Tissues; Transcript; Tyrosine Phosphorylation; Viral; Virus; Virus Diseases; cell type; knock-down; molecular domain; monocyte; mutant; novel; prevent; self-renewal; therapeutic target; C-terminal; CD4 Positive T Lymphocytes; Cells; Complex; Cyclins; Data; Dissection; Goals; HIV; HIV Infections; HIV-1; HIV-2; Hour; Infection; Knock-out; Macrophage; Mass Spectrum Analysis; Mediating; Molecular; Mutagenesis; Nucleotides; Pathway interactions; Phosphorylation; Phosphorylation Site; Phosphotransferases; Production; Proteins; Rest; SAM Domain; Specificity; Tissues; Transcript; Tyrosine Phosphorylation; Viral; Virus; Virus Diseases; cell type; knock-down; molecular domain; monocyte; mutant; novel; prevent; self-renewal; therapeutic target

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.

抽象的 巨噬细胞充当艾滋病毒储量，这是治愈的障碍。但是，分子机制 该病毒在这种细胞类型中的建立和维持感染的理解少得多。阐明 巨噬细胞中HIV复制的机制对于消除HIV的工作至关重要。 我们先前证明了Cyclin L2是巨噬细胞中HIV-1感染所必需的，与和相互作用 靶向HIV限制因子SAM结构域和含HD结构域的蛋白1（SAMHD1）进行降解。 但是，细胞周期蛋白L2-SAMHD1相互作用的细节尚不清楚。另外，如何调节细胞周期蛋白L2 在巨噬细胞中是未知的。在我们最近的研究中，我们表明Cyclin L2与磷酸化相互作用，并且是磷酸化的 通过双重特异性酪氨酸磷酸化调节激酶1a（dyrk1a）。而细胞周期蛋白L2的敲除 HIV感染减少了10倍，DYRK1A的耗竭增加了原发性巨噬细胞中HIV感染 10倍。该提案的总体目标是确定Cyclin L2，Dyrk1a之间的相互作用如何 SAMHD1调节巨噬细胞中的HIV复制。我们的具体目的是： AIM 1：定义激酶DyRK1A调节巨噬细胞中细胞周期蛋白L2的机制。我们 假设DYRK1A通过磷酸化或降解抑制细胞周期蛋白L2。首先，我们将确定 DYRK1A介导的细胞周期蛋白L2的磷酸化是否可以防止促进HIV感染 巨噬细胞。其次，我们将定义细胞周期蛋白L2和dyrk1a的分子结构域 互动。第三，我们将确定dyRK1a调节细胞周期蛋白L2水平的机制 巨噬细胞。 AIM 2：确定在病毒感染期间，细胞周期蛋白L2和HIV-1如何调节巨噬细胞中的SAMHD1。艾滋病病毒- 1没有VPX，但是尽管SAMHD1丰富，但仍能够在巨噬细胞中建立感染。 因此，HIV可以通过Cyclin L2或其他方式来策划SAMHD1的降解。在这个目标中，我们 （i）确定这种相互作用所需的细胞周期蛋白L2和SAMHD1的分子结构域（ii）确定 病毒或细胞因子（s）分别诱导细胞周期蛋白L2和SAMHD1水平升高和降低。 早期的HIV感染和（iii）确定巨噬细胞中的SAMHD1降解是否受细胞周期蛋白L2的影响 DyRK1a的磷酸化。 这些研究将揭示细胞周期蛋白L2-DYRK1A-SAMHD1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条确定艾滋病毒复制的新途径将非常重要 在巨噬细胞中受到限制或启用。潜在影响很大，因为这些蛋白质的解剖 相互作用可以揭示针对HIV感染的潜在治疗靶标。