Project 6: Vpr subversion of DNA repair

项目6：Vpr颠覆DNA修复

• 批准号: 9407944
• 负责人: jinwoo ahn
• 金额: $ 47.12万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/9407944
• 关键词: Biochemical; Biophysics; Cell Cycle Arrest; Chromatin; Complement; Complementary DNA; Complex; DNA; DNA Damage; DNA Repair; DNA Repair Gene; DNA Repair Pathway; DNA Structure; DNA biosynthesis; DNA damage checkpoint; DNA replication fork; Data; Disadvantaged; Ensure; Enzymes; Event; Excision; Excision Repair; Generations; HIV; HIV-1; HIV-2; Life Cycle Stages; Link; Mediating; Mitosis; NMR Spectroscopy; Pathway interactions; Phenotype; Primate Lentiviruses; Process; Proteins; Proteomics; Recruitment Activity; Reverse Transcription; Ribonucleotides; SMARCA3 gene; Site; Structure; Surgical Flaps; T-Lymphocyte; Testing; Uncertainty; Uracil; Viral Proteins; Virus; X-Ray Crystallography; homologous recombination; protein activation; protein complex; ubiquitin-protein ligase; vpr Gene Products

P6. Abstract The accessory protein Vpr facilitates HIV-1 replication in dividing T cells, ensuring orderly progression through the HIV-1 life cycle. The most remarkable Vpr phenotype is induction of DNA damage checkpoint and cell cycle arrest in G2/M phase, although the underlying details are the still elusive. There is compelling evidence that Vpr interacts with several cellular proteins that recognize damaged DNA, targeting them for proteasomal degradation via CRL4DCAF1 E3 ubiquitin ligase. While evident that, in principle, Vpr antagonism of DNA repair ultimately can be of benefit to HIV-1, the full extent of Vpr-mediated subversion of cellular DNA repair is not known. In this project, we propose to systematically explore the extent of Vpr's engagement with the DNA repair machinery, focusing on DNA repair pathways that recognize and process “marks of damage”, introduced into HIV-1 cDNA during reverse transcription. We aim to discover and validate other specific DNA repair protein(s) that are targeted by Vpr, in particular those mediating the induction of DNA damage checkpoint and cell cycle arrest. Further, we will biochemically, biophysically and structurally analyze the interactions between Vpr and the already identified targets HLTF, MUS81-EME1, and hHR23A, as well as any new target(s) discovered in this project. Overall, our studies will define the extent of Vpr-induced changes in cellular DNA repair, identify specific DNA repair pathways/proteins targeted by Vpr, and structurally characterize the responsible CRL4DCAF1 E3/Vpr/target complexes.

P6.摘要 辅助蛋白 Vpr 促进分裂 T 细胞中 HIV-1 的复制，确保有序进展 HIV-1生命周期最显着的Vpr表型是DNA损伤检查点和细胞周期的诱导。 尽管基本细节仍然难以捉摸，但有令人信服的证据表明 Vpr 已在 G2/M 阶段被捕。 与识别受损 DNA 的多种细胞蛋白相互作用，靶向它们进行蛋白酶体降解 通过CRL4DCAF1 E3泛素连接酶虽然明显表明，原则上Vpr最终可以拮抗DNA修复。 虽然 Vpr 介导的细胞 DNA 修复破坏的全部程度尚不清楚，但它对 HIV-1 有益。 项目中，我们建议系统地探索 Vpr 与 DNA 修复机制的参与程度， 专注于识别和处理引入 HIV-1 cDNA 的“损伤标记”的 DNA 修复途径 我们的目标是发现并验证其他特定的 DNA 修复蛋白。 Vpr 的靶向，特别是那些介导 DNA 损伤检查点和细胞周期停滞诱导的药物。 此外，我们将从生化、生物物理和结构上分析 Vpr 和 已经确定的目标 HLTF、MUS81-EME1 和 hHR23A，以及在此发现的任何新目标 总体而言，我们的研究将确定 Vpr 诱导的细胞 DNA 修复变化的程度，确定具体的情况。 Vpr 靶向的 DNA 修复途径/蛋白质，并在结构上表征了负责的 CRL4DCAF1 E3/Vpr/目标复合物。