Viral Chromatin Methylation as a Host Defense Against Geminiviruses

智力优势。该项目关注植物宿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个被称为双生病毒的含DNA植物病毒家族。 在植物中，RNA沉默是一种重要的抗病毒防御，导致病毒mRNA的特异性降解。在DNA病毒的情况下，一个相关的过程（RNA指导的甲基化）导致病毒DNA和相关的组蛋白的甲基化，这些组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，用于甲基化内源性细胞基因和病毒DNA的细胞途径基本相同。本项目将以双生病毒为模型进一步研究RNA介导的甲基化途径。具体而言，细胞RNA聚合酶II，IV和V在病毒基因组从头甲基化的作用将使用遗传和生物化学方法的组合进行检查。 将研究将这些酶募集到病毒染色体的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鉴定和表征识别染色质上甲基化标记的细胞蛋白，并将其转化为基因表达的变化。甲基化在正常发育期间控制细胞基因表达，并且对于组织细胞染色体的非活性区域和沉默潜在破坏性的侵入性DNA是重要的。因此，研究双生病毒染色质甲基化有望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。对宿主抗病毒防御的更好理解将最终导致能够更好地抵抗或抵抗病毒攻击的植物的发展。该项目将通过促进培训访问科学家、博士后研究员、研究生和本科生（其中一些人是代表性不足的少数民族），将研究和教育结合起来。此外，PI还参加了由俄亥俄州州立大学分子遗传学系赞助的NSF REU计划，以及OSU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的基本技术和原理培训。PI还通过向即将启动的新植物抗体设施捐赠抗体做出了重大贡献。作为过去16年来分子，细胞和发育生物学（MCDB，目前约130名博士生）的校园跨学科研究生课程的主任，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计划和多样性和包容性的大学办公室招募少数民族学生的积极作用。PI还积极关注教师和学生之间的互动，每年共同组织MCDB研讨会系列，跨学科研究生课程研讨会和第一年的研究生研讨会，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.