Viral Chromatin Methylation as a Host Defense Against Geminiviruses

智力上的功绩。该项目涉及植物寄主识别病毒病原体并启动特定防御的机制。这项研究的重点是一种被称为双生病毒的含有DNA的植物病毒家族。在植物中，RNA沉默是一种重要的抗病毒防御机制，可以导致病毒mRNA的特异性降解。对于DNA病毒，一个相关的过程(RNA导向的甲基化)导致病毒DNA和相关组蛋白的甲基化，这些蛋白共同构成病毒染色体。甲基化通常会导致基因沉默，该实验室最近的工作表明，用于甲基化内源细胞基因的细胞路径和病毒DNA本质上是相同的。在这个项目中，双生病毒将被用作进一步研究RNA引导的甲基化途径的模型。具体地说，细胞RNA聚合酶II、IV和V在病毒基因组从头甲基化中的作用将使用遗传和生化方法相结合的方法进行研究。我们将研究将这些酶招募到病毒染色体上的DNA序列特征，它们识别的序列的甲基化状态，以及它们产生的非编码RNA的性质。此外，双生病毒系统将用于识别和表征细胞蛋白质，这些蛋白质识别染色质上的甲基化标记，并将这些标记转化为基因表达的变化。甲基化在正常发育过程中控制细胞基因的表达，对于组织细胞染色体的非活跃区和沉默潜在的破坏性入侵DNA是重要的。因此，研究双生病毒染色质甲基化有助于增加我们对病毒发病机制和控制细胞染色质活性的基本过程的理解。更好地了解宿主的抗病毒防御最终将导致开发出能够更好地抵御或抵抗病毒攻击的植物。该项目将把研究和教育结合起来，帮助培训来访的科学家、博士后助理、研究生和本科生，他们中的一些人是代表不足的少数民族。此外，PI还参加了由俄亥俄州立大学分子遗传学系赞助的NSF REU计划，以及俄亥俄州立大学植物功能基因组学夏季实践研讨会，该研讨会吸引了国际和国内学生、博士后、年轻教师和高中教师，并对他们进行分子生物学和基因组学基本技术和原理的培训。PI还通过向即将推出的新的植物抗体设施捐赠抗体而做出了实质性贡献。在过去的16年里，作为分子、细胞和发育生物学(MCDB)全校跨学科研究生项目(MCDB，目前约有130名博士生)的主任，PI在促进研究生教育方面有着悠久的历史。努力包括通过研究生院的SROP计划和大学多样性和包容性办公室在招收少数族裔学生方面发挥积极作用。PI还对促进师生之间的互动非常感兴趣，并每年联合举办MCDB研讨会系列、跨学科研究生项目研讨会和研究生一年级研讨会，其中包括研究伦理培训。

Intellectual Merit. This project concerns mechanisms by which plant hosts recognize viral pathogens and initiate specific defenses. The research is focused on a family of DNA-containing plant viruses known as geminiviruses. In plants, RNA silencing is an important antiviral defense that results in specific degradation of viral mRNA. In the case of DNA viruses, a related process (RNA-directed methylation) leads to the methylation of viral DNA and associated histone proteins that together constitute the viral chromosome. Methylation typically leads to gene silencing, and recent work from this laboratory has demonstrated that cellular pathways used to methylate endogenous cellular genes and viral DNA are essentially identical. In this project, geminiviruses will be employed as models to further study the RNA-directed methylation pathway. Specifically, the roles of cellular RNA polymerases II, IV, and V in de novo methylation of viral genomes will be examined using a combination of genetic and biochemical approaches. The DNA sequence features that recruit these enzymes to the viral chromosome, the methylation status of the sequences that they recognize, and the nature of the non-coding RNAs that they generate will be investigated. In addition, the geminivirus system will be used to identify and characterize cellular proteins that recognize methylation marks on chromatin and translate these into changes in gene expression. Methylation controls cellular gene expression during normal development, and is important for organizing inactive regions of cellular chromosomes and silencing potentially damaging invasive DNAs. Thus studying geminivirus chromatin methylation promises to increase our understanding of viral pathogenesis and of fundamental processes that govern the activity of cellular chromatin.Broader Impacts. A better understanding of host antiviral defenses will eventually lead to the development of plants that are better able to withstand or resist virus attack. This project will integrate research and education by contributing to the training of visiting scientists, postdoctoral associates, graduate students, and undergraduate students, some of whom are underrepresented minorities. In addition, the PI participates in the NSF REU Program sponsored by the Department of Molecular Genetics at Ohio State University, and in the OSU Summer Practical Workshop in Plant Functional Genomics, which attracts international and domestic students, postdocs, young faculty, and high school teachers and trains them in basic techniques and principles of molecular biology and genomics. The PI has also contributed materially by donating antibodies to a new, soon-to-be-launched Plant Antibody Facility. As Director of the campus-wide interdisciplinary graduate program in Molecular, Cellular, and Developmental Biology (MCDB, currently ~130 PhD students) for the past 16 years, the PI has a long history of promoting graduate education. Efforts include an active role in recruiting minority students through the Graduate School's SROP Program and the University Office of Diversity and Inclusion. The PI also has an active interest in enabling interactions between faculty and students and annually co-organizes the MCDB Seminar Series, the Interdisciplinary Graduate Program Symposium, and the First-Year Graduate Student Seminar, which includes training in research ethics.