Structural Biophysics of HIV Splicing and Transcriptional Control

HIV剪接和转录控制的结构生物物理学

• 批准号: 10917782
• 负责人: Blanton Smith Tolbert
• 金额: $ 54.86万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-08-19 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10917782
• 关键词: 3-Dimensional; Acquired Immunodeficiency Syndrome; Adopted; Binding; Binding Sites; Biochemical; Biological Assay; Biophysics; Cells; Chemicals; Complex; Cytoplasm; Data; Elements; Etiology; Event; Foundations; Gene Expression; Genetic Transcription; HIV; Health; Heterogeneous-Nuclear Ribonucleoproteins; Human; Image; Immunofluorescence Immunologic; Knowledge; Maps; Mediating; Modeling; Molecular; Molecular Conformation; Mutation; Nature; Nuclear; Pathway interactions; Phenotype; Positive Transcriptional Elongation Factor B; Process; Property; Proteins; RNA; RNA Splicing; RNA-Binding Proteins; RNA-Protein Interaction; Regulatory Element; Research; Series; Site; Small Nuclear RNA; Specificity; Structural Biochemistry; Structural Models; Structure; Tars; Techniques; Testing; Therapeutic Intervention; Transcript; Transcription Elongation; Transcriptional Elongation Factors; Transcriptional Regulation; Transcriptional Silencer Elements; Untranslated RNA; Viral; Viral Pathogenesis; Virus; Virus Latency; Virus Replication; antiretroviral therapy; base; biophysical properties; hnRNP protein A1; inhibitor; innovation; insight; new therapeutic target; novel; programs; protein complex; protein protein interaction; reactivation from latency; spatiotemporal; success; three dimensional structure; viral RNA

Project abstract: Despite advances in antiretroviral therapy, HIV continues to pose serious threats to human health and global economies. Knowledge gaps in understanding the basic mechanisms that control HIV gene expression present critical barriers to identifying novel targets for therapeutic intervention or strategies to coun- teract viral latency. In this proposal, we endeavor to close those gaps concerning the contributions of RNA struc- tural dynamics to HIV splicing and transcriptional control. We posit that HIV uses structured RNA elements to direct the cooperative assembly of specific RNA-protein (RNP) complexes that in turn modulates splicing. This type of RNA-mediated cooperativity is an innovative concept that explains why HIV conserves RNA structures near its splice sites. Those structures and its conformational dynamics can either positively or negatively impart cooperative protein-protein interactions; we seek to understand the biophysical nature of such mechanisms. We present strong evidence that the RNA interactions surrounding HIV splice sites A2 and A3 intrinsically regulate the levels of vpr and tat, respectively. Mutations within those RNA structures inhibit HIV replication by deregu- lating RNA splicing. We further contend that similar RNA-mediated cooperative mechanisms determine the ex- tent to which the cellular P-TEFb is released from 7SK snRNA and made available to stimulate HIV transcription elongation. To test the extent to which RNA-mediated cooperativity influences HIV nuclear gene expression, we have developed a technically innovative research program that integrates cutting edge biophysical and biochem- ical techniques to extract mechanistic principles about host-virus RNP complexes. We will implement this pro- gram by adopting two broadly conceptual specific aims: 1. Molecular interactions of viral RNAs and RNPs that contribute to HIV splicing and 2. Molecular interactions of 7SK snRNA and its different RNPs that contribute to HIV transcription. The success of this proposal promises to deliver unprecedented in-sights into the structural dynamics of RNA-RNA and protein-RNA interactions that control HIV gene expression and it may inform on details into molecular mechanisms that contribute to latency. This will in turn provide a foundation for identifying novel targets for cure strategies.

项目摘要：尽管抗逆转录病毒疗法取得了进展，但艾滋病毒仍对人类构成严重威胁 健康和全球经济。在了解控制HIV基因的基本机制方面的知识差距 表达是确定新的治疗干预靶点或应对策略的关键障碍。 特技病毒潜伏期。在这项建议中，我们努力弥合这些关于RNA结构的贡献的空白。 艾滋病毒剪接和转录控制的文化动态。我们假设HIV使用结构化的RNA元件来 指导特定RNA-蛋白质(RNP)复合体的合作组装，进而调节剪接。这 RNA介导的协作性类型是一个创新的概念，它解释了为什么HIV保留了RNA结构 在它的拼接点附近。这些结构及其构象动力学可以是积极的，也可以是消极的 蛋白质之间的协作相互作用；我们试图了解这种机制的生物物理性质。我们 提供强有力的证据表明，HIV剪接位点A2和A3周围的RNA相互作用本质上调节 分别测定VPR和TAT水平。这些RNA结构中的突变通过去突变抑制HIV复制- 迟来的RNA剪接。我们进一步认为，类似的RNA介导的合作机制决定了前两者之间的关系。 从7SK SnRNA中释放细胞内P-TEFb并用于刺激HIV转录的帐篷 伸长率。为了测试RNA介导的协作性对HIV核基因表达的影响程度，我们 开发了集尖端生物物理和生物化学为一体的技术创新研究计划- 提取宿主-病毒RNP复合体的机制原理的技术。我们将实施这一政策- 通过采用两个概念性的具体目标：1.病毒RNA和RNPs的分子相互作用 有助于HIV的剪接和2.7SK SnRNA及其不同RNPs的分子相互作用 艾滋病毒转录。这项提案的成功有望对结构性问题提供前所未有的洞察力 控制HIV基因表达的RNA-RNA和蛋白质-RNA相互作用的动力学及其可能提供的信息 详细了解导致潜伏期的分子机制。这将反过来为确定 治疗策略的新靶点。

项目成果

R-BIND 2.0: An Updated Database of Bioactive RNA-Targeting Small Molecules and Associated RNA Secondary Structures.

• DOI: 10.1021/acschembio.2c00224
• 发表时间: 2022-06-17
• 期刊: ACS CHEMICAL BIOLOGY
• 影响因子: 4
• 作者: Donlic, Anita;Swanson, Emily G.;Chiu, Liang-Yuan;Wicks, Sarah L.;Juru, Aline Umuhire;Cai, Zhengguo;Kassam, Kamillah;Laudeman, Chris;Sanaba, Bilva G.;Sugarman, Andrew;Han, Eunseong;Tolbert, Blanton S.;Hargrove, Amanda E.
• 通讯作者: Hargrove, Amanda E.

Biophysical Analysis of Small Molecule Binding to Viral RNA Structures.

小分子与病毒 RNA 结构结合的生物物理分析。

• DOI: 10.1007/978-1-0716-2695-5_16
• 发表时间: 2023
• 期刊: Methods in molecular biology (Clifton, N.J.)
• 作者: Chiu,Liang-Yuan;Davila-Calderon,Jesse;Cai,Zhengguo;Tolbert,BlantonS
• 通讯作者: Tolbert,BlantonS

Encoded Conformational Dynamics of the HIV Splice Site A3 Regulatory Locus: Implications for Differential Binding of hnRNP Splicing Auxiliary Factors.

• DOI: 10.1016/j.jmb.2022.167728
• 发表时间: 2022-09-30
• 期刊: JOURNAL OF MOLECULAR BIOLOGY
• 影响因子: 5.6
• 作者: Chiu, Liang-Yuan;Emery, Ann;Jain, Niyati;Sugarman, Andrew;Kendrick, Nashea;Luo, Le;Ford, William;Swanstrom, Ronald;Tolbert, Blanton S.
• 通讯作者: Tolbert, Blanton S.

HnRNP A1/A2 Proteins Assemble onto 7SK snRNA via Context Dependent Interactions.

HNRNP A1/A2蛋白通过上下文依赖性相互作用聚集在7SK snRNA上。

• DOI: 10.1016/j.jmb.2021.166885
• 发表时间: 2021-04-30
• 期刊: Journal of molecular biology
• 影响因子: 5.6
• 作者: Luo L;Chiu LY;Sugarman A;Gupta P;Rouskin S;Tolbert BS
• 通讯作者: Tolbert BS

The Repurposing of Cellular Proteins during Enterovirus A71 Infection.

• DOI: 10.3390/v16010075
• 发表时间: 2023-12-31
• 期刊: Viruses
• 作者: Abedeera SM;Davila-Calderon J;Haddad C;Henry B;King J;Penumutchu S;Tolbert BS
• 通讯作者: Tolbert BS