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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.

SERINC 5（Ser 5）是一种新型的限制性因子，可强烈阻断HIV-1进入。Ser 5属于丝氨酸 SERINC（SERINC）家族，有5个成员（1到5）。它们是III型整合膜蛋白， 9-11跨膜结构域。Ser 5和Ser 3最初被确定为HIV-1 Nef的对抗靶点增加病毒的传染性虽然SERINC蛋白具有31-58%的序列同源性，但只有Ser 5和Ser 6具有相同的序列同源性。 Ser 3具有抗病毒活性，尽管Ser 3活性较弱。此外，虽然Ser 4的抗病毒活性是据报道，它在蛋白质水平上表达很差。Ser 5被HIV-1 Nef以及MLV glycoGag拮抗 EIAV S2。Ser 5的Nef拮抗作用在灵长类慢病毒的流行中起重要作用，他们的主人。因此，Ser 5是逆转录病毒的重要限制因子。我们的目标是研究 Ser 5如何抑制HIV-1复制的机制，以及Ser 5如何被Nef抵消。一是将阐明Ser 5如何阻止HIV-1进入。Ser 5被包装成病毒体以阻止病毒进入，但其机制对目前仍不清楚.还不清楚为什么Ser 5抑制依赖于病毒Env糖蛋白。1级病毒，主要是实验室适应的病毒，非常敏感，而大多数2/3级病毒，这是巨大的大多数循环菌株对Ser 5有抗性。值得注意的是，天然第1层Env三聚体主要占据了开放构象，而那些2/3层三聚体占据闭合构象。此外，Ser 5使艾滋病毒- 1变得对中和抗体敏感，这表明Ser 5可以修饰Env构象。我们假设Ser 5在开放状态下破坏Env三聚体，导致Env失活和HIV-1的阻断入境我们将使用灵敏的双分子荧光互补（BiFC）检测来研究Env 寡聚化和SERINC-Env相互作用，并使用病毒学试验阐明HIV-1如何进入被丝氨酸5阻断，也可能被丝氨酸4阻断。其次，我们将阐明Nef如何拮抗Ser 5。NEF 通过下调质膜上的Ser 5并将其排除在病毒体之外来抵消Ser 5，但精确的机制尚不清楚。我们报道Nef通过受体介导的内吞作用快速内化Ser 5，并将Ser 5重新定位于内体。然后丝氨酸5以泛素化依赖的方式靶向溶酶体用于降解。细胞周期蛋白K（Cyclin K，CycK）是一种Nef结合蛋白，与细胞周期蛋白依赖性激酶结合 (CDK)12和13中所示的那些，并且调节基因表达和基因组稳定性。我们发现CycK和CDK 13是需要Nef下调Ser 5。我们假设Nef将CDK 13/CycK募集到质膜上，使Ser 5磷酸化并将Ser 5靶向至内体/溶酶体途径进行降解。我们会研究通过检测Ser 5磷酸化和CDK 13/CycK复合物在Nef下调Ser 5中的关键作用，确定磷酸化如何影响Ser 5细胞内运输和/或降解。新的抗逆转录病毒药物机制和/或目标将从我们的研究中发现，这可能会转化为新的抗逆转录病毒疗法