Mechanism of Viral Genome Delivery into Cells

病毒基因组传递至细胞的机制

• 批准号: 10160588
• 负责人: Gino Cingolani
• 金额: $ 34.31万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-06-01 至 2026-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10160588
• 关键词: Bacteria; Bacteriophages; Biological; Biological Process; Biology; Biotechnology; Cell Nucleus; Cells; Complex; Cryoelectron Microscopy; DNA; Development; Eukaryotic Cell; Funding Mechanisms; Genome; Gram-Negative Bacteria; Guanosine Triphosphate Phosphohydrolases; Herpesviridae; Human; Immune system; Importins; Injections; Laboratories; Literature; Mediating; Medicine; Membrane; Membrane Proteins; Molecular; Molecular Machines; Nuclear Import; Nuclear Pore Complex; Organelles; Principal Investigator; Process; Protein Biochemistry; Publications; Regulation; Research; Science; Signal Transduction; Structure; Viral; Viral Genome; Virulence; Virus; Work; X-Ray Crystallography; cell envelope; genetic information; interest; macromolecule; nanomachine; novel therapeutic intervention; nucleocytoplasmic transport; pressure; programs; receptor; structural biology

Project Summary Nearly 120 years since the discovery of the first virus, our understanding of how viruses deliver genomes into cells overcoming the complexity of biological membranes remains limited. While a vast scientific literature exists on viral surface proteins and their interaction with host receptors, and the immune system, little emphasis has been devoted to studying the delivery of entire viral genomes into cells. For instance, how do bacteriophages eject DNA through the cell envelope of Gram-negative bacteria? Or, in humans, how do herpesviruses deliver ~200 kb genome through the Nuclear Pore Complex (NPC) into the cell nucleus? For a quarter of a century, first as a trainee (1995-2003), and since 2004 as a principal investigator, I have investigated the mechanisms of nucleocytoplasmic transport and viral genome packaging. My work has resulted in close to 85 publications that contributed to elucidating the atomic structure and regulation of crucial factors implicated in nuclear import, and viral genome packaging. In this R35, I propose to combine the study of these two seemingly distinct biological processes by focusing on the mechanisms of viral genome delivery into living cells. Specifically, I will ask two biological questions that seek to compare and contrast how simple bacterial viruses (or bacteriophages) eject their DNA into bacteria with how Herpesviruses deliver their complex genomes into the nucleus of eukaryotic cells. The first question explores how bacteriophages eject ~45 kb genomes through the cell envelope of gram-negative bacteria. Long-thought to be a simple pressure-driven injection, this process uses a virus-encoded nanomachine, which we have begun to study in my laboratory. The second question explores how Herpesviruses deliver their large genome through the Nuclear Pore Complex (NPC) of human cells into the cell nucleus. This is a signal- and energy-mediated process that uses host importins and the GTPase Ran, exploiting the cellular transport machinery to promote entry of an exogenous genome into the nucleus. Overall, understanding how viruses transfer genetic information through biological membranes into cells and organelles is vital for deciphering the molecular mechanisms of virulence as well as the development of novel therapeutic approaches. The common denominator of this R35 lies in our interest in the structure and transport mechanisms of biological macromolecules. Our research approach marries established sciences like protein biochemistry and X-ray crystallography with the power of cryo-electron microscopy (cryo-EM) to visualize biological macromolecules in near-native conditions. We believe that this R35 MIRA funding mechanism will fuel the creative and diligent pursuit of answers to the questions we pose, permitting our research program to achieve significant advancements in structural biology.

项目摘要 自发现第一种病毒以来，将近120年，我们对病毒如何将基因组传递到 克服生物膜复杂性的细胞仍然有限。而庞大的科学文学 存在于病毒表面蛋白上及其与宿主受体的相互作用，免疫系统很少 重点已致力于研究整个病毒基因组向细胞的递送。例如，如何 噬菌体通过革兰氏阴性细菌的细胞包膜排出DNA？或者，在人类中如何 疱疹病毒通过核孔复合物（NPC）传递约200 kb的基因组进入细胞核？ 四分之一世纪，首先是一名学员（1995-2003），自2004年以来，我就有 研究了核质运输和病毒基因组包装的机制。我的工作有 导致近85个出版物有助于阐明原子结构和监管 涉及核进口和病毒基因组包装的因素。在此R35中，我建议将研究结合起来 通过关注病毒基因组传递的机制，这两个看似不同的生物过程 进入活细胞。具体而言，我将提出两个生物学问题，以比较和对比多么简单 细菌病毒（或噬菌体）将其DNA驱射到细菌中，疱疹病毒如何提供其 复杂的基因组进入真核细胞的核。第一个问题探讨了噬菌体如何 通过革兰氏阴性细菌的细胞膜喷射〜45 Kb基因组。长期思考是一个简单的 压力驱动的注射，此过程使用病毒编码的纳米机械，我们已经开始研究 我的实验室。第二个问题探讨了疱疹病毒如何通过 人类细胞的核孔复合物（NPC）进入细胞核。这是一个信号和能量介导的 使用宿主导入蛋白和GTPase运行的过程，利用蜂窝传输机械来促进 外源基因组进入细胞核。总体而言，了解病毒如何转移遗传 通过生物膜进入细胞和细胞器的信息对于破译分子至关重要 毒力的机制以及新型治疗方法的发展。共同 该R35的分母在于我们对生物学的结构和运输机制的兴趣 大分子。我们的研究方法与蛋白质生物化学和X射线等科学结合 晶体学具有冷冻电子显微镜（Cryo-EM）的力量，可视化生物大分子中的生物大分子 近乎本地的条件。我们认为，这种R35 MIRA资金机制将助长创造力和勤奋 追求我们提出的问题的答案，允许我们的研究计划获得重要的 结构生物学的进步。