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Mechanistic role of ns2 protein in evasion of innate immune response by alphaviruses

ns2蛋白在甲病毒逃避先天免疫反应中的机制作用

基本信息

批准号：
10165475
负责人：
ELENA I FROLOVA
金额：
$ 60.36万
依托单位：
UNIVERSITY OF ALABAMA AT BIRMINGHAM
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-06-16 至 2023-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10165475
关键词：
Africa Alphavirus Animals Antarctic Antiviral Response Arthritogenic Attenuated Binding Blood Circulation Catalytic Domain Cause of Death Cells Central America Chikungunya virus Complex Cytoplasm DNA DNA-Directed RNA Polymerase Data Degradation Pathway Development Disease Outbreaks Dissection Encephalitis Encephalitis Viruses Equine Encephalomyelitis Family Florida Genetic Transcription Goals Health Hour Human Immune Evasion Immunity Immunologic Deficiency Syndromes India Infection Innate Immune Response Integration Host Factors Interferons Investigation Island Italy Mediating Methyltransferase Molecular Mutate Mutation Neurologic Nonstructural Protein Nuclear Nucleic Acid Binding Oceania Outcome Pathogenesis Pathogenicity Play Proteins RNA RNA Polymerase II RNA replication Research Research Proposals Risk Role S-Adenosylmethionine Safety Signal Transduction South America Surface Testing Time Togaviridae Transcription-Coupled Repair Travel Vaccines Variant Venezuelan Venezuelan Equine Encephalitis Virus Virus Virus Replication West Indies Western Equine Encephalitis Virus base chikungunya infection forest helicase human disease improved innate immune mechanisms mutant neglect novel therapeutics novel vaccines pathogen prevent therapeutic development vaccine candidate virus host interaction

项目摘要

Alphaviruses are an important group of human and animal pathogens that are widely distributed on all continents. Until recently, the importance of the Old World alphaviruses in global human health was strongly underestimated. However, recent outbreaks of chikungunya virus (CHIKV) in India, Italy, East Africa's coastal islands and most recently on the Caribbean islands, South and Central America, Oceania have highlighted the risk posed by this virus. Our previous studies have unambiguously demonstrated that the Old World alphaviruses employ their nonstructural protein nsP2 to re-direct the cellular transcription coupled repair pathway for degradation of the catalytic subunit of the DNA- dependent RNA polymerase II, RPB1, and thus, to completely inhibit cellular transcription. Within a few hours post infection, virus-induced transcriptional shutoff makes the cells incapable of initiating an antiviral response and leads to development of cytopathic effect. Thus, the Old World alphavirus nsP2 is the major determinant of pathogenesis at the molecular and cellular levels. We have also demonstrated that the transcription inhibitory functions of nsP2 are determined not by its proteolytic activity, but by the ability of this protein to function as a helicase, and by its S-adenosyl-L- methionine-dependent RNA methyltransferase-like (SAM-like) domain. Two specific aims of the proposed research plan are focused on i) mechanistic understanding of the nsP2 helicase domain function in stalling cellular DNA-dependent RNA polymerase II and ii) on identification of cellular proteins that interact with the SAM-like domain of nsP2. We will also define the mechanistic role of the interactions of the latter domain in transcription inhibition. In our preliminary studies, we have developed a set of CHIKV variants with mutated nsP2, which are incapable of interfering with development of the innate immune response to CHIKV replication. These irreversible mutations had no detectable effect on the rates of CHIKV replication in cells defective in type I IFN signaling, but are cleared from IFN-competent cells without development of cytopathic effect. Therefore, in the third specific aim, we intend to assess the potential of these nsP2 mutants to improve the safety of the currently available experimental vaccine CHIKV 181/25.

甲型病毒是一组重要的人类和动物病原体，分布广泛。分布于各大洲。直到最近，旧大陆甲型病毒的重要性对全球人类健康的影响被严重低估。然而，最近爆发的霍乱印度、意大利、东非沿海岛屿和最近发现的基孔肯雅病毒(CHIKV) 在加勒比海岛屿、南美洲和中美洲，大洋洲强调了这种风险由这种病毒引起的。我们之前的研究已经明确地证明，旧大陆甲型病毒利用它们的非结构蛋白nsP2重定向细胞 DNA催化亚基降解的转录偶联修复途径依赖于RNA聚合酶II，RPB1，从而完全抑制细胞抄写。在感染后的几个小时内，病毒诱导的转录关闭使细胞不能启动抗病毒反应，并导致细胞病变效应。因此，东半球甲型病毒nsP2是在分子和细胞水平上的发病机制。我们还证明了 NsP2的转录抑制功能不是由它的蛋白分解活性决定的，而是由它的活性决定的。通过这种蛋白质作为解旋酶的能力，以及通过它的S-腺苷-L- 蛋氨酸依赖的RNA甲基转移酶样(SAM-like)结构域。两个具体的拟议研究计划的目标集中在：1)机械地理解 NsP2解旋酶结构域在细胞DNA依赖的RNA聚合酶II停滞中的作用以及ii)鉴定与SAM样结构域相互作用的细胞蛋白 NsP2。我们还将定义后一个领域的交互在转录抑制。在我们的初步研究中，我们已经开发了一套CHIKV 具有突变的nsP2的变体，这些突变不能干扰对CHIKV复制的先天免疫反应。这些不可逆转的突变没有 I型干扰素缺陷细胞中CHIKV复制率的可检测效应信号，但从干扰素活性细胞中清除，不会发生细胞病变效果。因此，在第三个具体目标中，我们打算评估这些 NsP2突变体以提高目前可用的实验疫苗的安全性 CHIKV 181/25。

项目成果

期刊论文数量（16）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Natural and Recombinant SARS-CoV-2 Isolates Rapidly Evolve In Vitro to Higher Infectivity through More Efficient Binding to Heparan Sulfate and Reduced S1/S2 Cleavage.

DOI：
10.1128/jvi.01357-21
发表时间：
2021-10-13
期刊：
Journal of virology
影响因子：
5.4
作者：
Shiliaev N;Lukash T;Palchevska O;Crossman DK;Green TJ;Crowley MR;Frolova EI;Frolov I
通讯作者：
Frolov I

All Domains of SARS-CoV-2 nsp1 Determine Translational Shutoff and Cytotoxicity of the Protein.

DOI：
10.1128/jvi.01865-22
发表时间：
2023-03-30
期刊：
JOURNAL OF VIROLOGY
影响因子：
5.4
作者：
Frolov, Ilya;Agback, Tatiana;Palchevska, Oksana;Dominguez, Francisco;Lomzov, Alexander;Agback, Peter;Frolova, Elena I.
通讯作者：
Frolova, Elena I.

Acquisition of Furin Cleavage Site and Further SARS-CoV-2 Evolution Change the Mechanisms of Viral Entry, Infection Spread, and Cell Signaling.