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The role of SERINC5 in HIV-1 replication

SERINC5 在 HIV-1 复制中的作用

基本信息

批准号：
10817137
负责人：
YONG-HUI ZHENG
金额：
$ 39.12万
依托单位：
UNIVERSITY OF ILLINOIS AT CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-07-08 至 2024-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10817137
关键词：
Adaptor Signaling Protein Affect Alternative Splicing Anabolism Anti-Retroviral Agents Binding Binding Proteins Biological Assay Cell membrane Cells Complement Complex Cyclin-Dependent Kinases Cyclins Degradation Pathway Destinations Development Down-Regulation Drug Targeting Endocytosis Endosomes Equine Infectious Anemia Virus Exclusion Exhibits Family Fluorescence Gene Expression Genome Stability Goals HIV Envelope Protein gp120 HIV-1 Human Infection Integral Membrane Protein Investigation Laboratories Lysosomes Mediating Membrane Fusion Molecular Molecular Conformation Murine leukemia virus Outcome Pathway interactions Phosphatidylserines Phosphorylation Play Prevalence Primate Lentiviruses Protein Isoforms Proteins Public Health Recycling Reporting Research Resistance Retroviridae Role Sequence Homology Serine Sorting Sphingolipids Testing Translating Transmembrane Domain Ubiquitination Viral Viral Physiology Viral Proteins Virion Virus Virus Replication Yeasts antagonist antiretroviral therapy env Gene Products env Glycoproteins member neutralizing antibody novel novel strategies pharmacologic receptor receptor binding receptor mediated endocytosis recruit screening trafficking viral entry inhibitor viral rescue yeast two hybrid system

项目摘要

SERINC5 (Ser5) is a novel restriction factor that strongly blocks HIV-1 entry. Ser5 belongs to the serine incorporator (SERINC) family that has five members (1 to 5). They are type III integral membrane proteins with 9-11 transmembrane domains. Ser5 and Ser3 were initially identified as the counteractive target of HIV-1 Nef that increases viral infectivity. Although SERINC proteins share 31-58% sequence homology, only Ser5 and Ser3 have antiviral activities, albeit the Ser3 activity is weak. In addition, although the Ser4 antiviral activity was reported, it is poorly expressed at protein levels. Ser5 is antagonized by HIV-1 Nef as well as MLV glycoGag and EIAV S2. The Nef antagonism of Ser5 plays an important role in the prevalence of primate lentiviruses in their hosts. Thus, Ser5 is an important restriction factor for retroviruses. Our goal is to investigate the molecular mechanisms of how Ser5 inhibits HIV-1 replication and conversely, how Ser5 is counteracted by Nef. First, we will elucidate how Ser5 blocks HIV-1 entry. Ser5 is packaged into virions to block viral entry, but the mechanism is still unclear. It is also unclear why the Ser5 inhibition depends on viral Env glycoproteins. Tier 1 viruses, which are largely laboratory-adapted viruses, are very sensitive, whereas most Tier 2/3 viruses, which are the vast majority of circulating strains, are resistant to Ser5. Notably, native Tier 1 Env trimers predominantly occupy an open conformation, whereas those Tier 2/3 trimers occupy a closed conformation. In addition, Ser5 makes HIV- 1 become sensitive to neutralizing antibodies, suggesting that Ser5 may modify the Env conformation. We hypothesize that Ser5 disrupts Env trimers in an open state, resulting in Env inactivation and blockade of HIV-1 entry. We will use a sensitive bimolecular fluorescence complementation (BiFC) assay to study Env oligomerization and SERINC-Env interactions in live cells and use virologic assays to elucidate how HIV-1 entry is blocked by Ser5 and also likely by Ser4. Second, We will elucidate how Nef antagonizes Ser5. Nef counteracts Ser5 by downregulating Ser5 from plasma membrane and excluding it from virions, but the precise mechanism is still unclear. We reported that Nef rapidly internalizes Ser5 via receptor-mediated endocytosis and re-localizes Ser5 to endosomes. Ser5 is then targeted to lysosomes in a ubiquitination-dependent manner for degradation. Cyclin K (CycK) is a previously identified Nef-binding protein that binds Cyclin-dependent kinase (CDK) 12 and 13 and regulates gene expression and genome stability. We found that CycK and CDK13 are required for Nef-downregulation of Ser5. We hypothesize that Nef recruits CDK13/CycK to plasma membrane to phosphorylate Ser5 and targets Ser5 to endosome/lysosome pathways for degradation. We will study the critical role of the CDK13/CycK complex in Nef downregulation of Ser5 by testing the Ser5 phosphorylation and determine how the phosphorylation affects the Ser5 intracellular trafficking and/or degradation. New antiretroviral mechanisms and/or targets will be discovered from our investigations, which is likely translated into novel antiretroviral therapies.

SERINC5(Ser5)是一种新的限制因子，可以强烈地阻止HIV-1的进入。Ser5属于丝氨酸有五个成员(1到5个)的合并者(SERINC)家族。它们是III型完整膜蛋白，具有 9-11个跨膜结构域。Ser5和Ser3最初被确定为HIV-1 Nef的对抗靶点这增加了病毒的传染性。尽管SERINC蛋白有31%-58%的序列同源性，但只有Ser5和 Ser3具有抗病毒活性，尽管Ser3的活性较弱。此外，尽管Ser4的抗病毒活性是据报道，它在蛋白质水平上表达很差。HIV-1Nef和MLV糖蛋白对Ser5的拮抗作用和EIAV S2。Ser5的Nef拮抗作用在灵长类慢病毒流行中起重要作用他们的主人。因此，Ser5是逆转录病毒的重要限制因子。我们的目标是研究分子 Ser5如何抑制HIV-1复制的机制，反之，Ser5如何被Nef抵消。首先，我们将阐明Ser5是如何阻止HIV-1进入的。Ser5被包装成病毒粒子以阻止病毒进入，但其机制目前仍不清楚。目前也不清楚为什么Ser5抑制依赖于病毒包膜糖蛋白。第1级病毒，其中主要是实验室适应的病毒，非常敏感，而大多数2/3级病毒，它们是巨大的大多数流通菌株对Ser5耐药。值得注意的是，本地Tier 1 Env Trimer主要占据开放构象，而那些Tier 2/3三聚体占据封闭构象。此外，Ser5使HIV- 1对中和抗体敏感，提示Ser5可能改变了Env的构象。我们假设Ser5在开放状态下干扰Env Trimer，导致Env灭活并阻断HIV-1 进入。我们将使用一种灵敏的双分子荧光互补(BIFC)分析来研究环境活细胞中的寡聚和SERINC-Env相互作用以及用病毒学方法阐明HIV-1是如何进入的被Ser5阻止，也可能被Ser4阻止。其次，我们将阐明Nef是如何对抗Ser5的。NEF 通过从质膜下调Ser5并将其排除在病毒粒子之外来中和Ser5，但精确的机制仍不清楚。我们报道Nef通过受体介导的内吞作用迅速内化Ser5 并将Ser5重新定位为内小体。然后，Ser5以泛素化依赖的方式靶向溶酶体为了降级。细胞周期蛋白K(Cyclin K，CycK)是一种与Nef结合的蛋白，可与细胞周期蛋白依赖性激酶结合 (CDK)12和13，并调节基因表达和基因组稳定性。我们发现CycK和CDK13是 Nef-下调Ser5所需的。我们假设Nef将CDK13/CycK招募到质膜上使Ser5磷酸化，并以Ser5为靶标，通过内切/溶酶体途径进行降解。我们会研究 CDK13/CycK复合体通过检测Ser5的磷酸化和转录水平在Nef下调Ser5中的关键作用确定磷酸化如何影响Ser5在细胞内的运输和/或降解。新型抗逆转录病毒药物机制和/或目标将从我们的调查中被发现，这很可能转化为小说抗逆转录病毒疗法。