CAREER: Dissecting the Mechanism of Replication Initiation in Vertebrates via Single Molecule Imaging

职业：通过单分子成像剖析脊椎动物复制起始机制

• 批准号: 2144481
• 负责人: Gheorghe Chistol
• 金额: $ 108.57万
• 依托单位: Stanford University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-01 至 2027-01-31
• 项目状态: 未结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2144481&HistoricalAwards=false
• 关键词: CAREER; Dissecting; Mechanism; Replication; Initiation

This award is funded in whole or in part under the American Rescue Plan Act of 2021 (Public Law 117-2). This project will employ a cutting-edge microscopy approach to investigate the earliest stage in DNA replication at a molecular level. Several proteins act in concert to copy DNA bidirectionally from sites called “replication origins”. How this process initiates bidirectionally from thousands of origins remains one of the most mysterious aspects of DNA biology. This research will advance basic understanding of DNA replication mechanisms that safeguard the integrity of genetic information. The educational goals of this project are: (1) to collaborate with teachers from a local underserved high school to develop a hands-on biotechnology curriculum, (2) to provide professional training opportunities for high-school science teachers and research opportunities for high-school students, and (3) to develop skills training courses for graduate students. American Rescue Plan funding provides support for this researcher at a critical stage in his career.Replication initiation involves short-lived interactions between DNA and numerous proteins, which are challenging to monitor and measure using current approaches. Although the process of replication initiation has been studied in detail in yeast, key proteins are poorly conserved between yeast and vertebrates and their exact functions remain unclear. This project will systematically dissect replication initiation in the Xenopus egg extract model system using novel single-molecule imaging methodology developed by the principal investigator. This effort will be complemented by biochemical, genomic, and proteomic approaches. The research has the potential to advance the field by (1) providing a quantitative view of replication initiation in higher eukaryotes, (2) identifying and characterizing unknown regulators of replication initiation, and (3) elucidating why key replication proteins have dramatically increased in complexity from yeast to humans.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

该奖项全部或部分根据2021年美国救援计划法案（公法117-2）资助。该项目将采用尖端的显微镜方法，在分子水平上研究DNA复制的最早阶段。几种蛋白质协同作用，从称为“复制起点”的位点双向复制DNA。这个过程如何从数千个来源双向启动仍然是DNA生物学最神秘的方面之一。这项研究将推进对DNA复制机制的基本理解，以保护遗传信息的完整性。该项目的教育目标是：（1）与当地一所服务水平低下的高中的教师合作，开发一门实践生物技术课程;（2）为高中科学教师提供专业培训机会，为高中学生提供研究机会;（3）为研究生开发技能培训课程。 美国救援计划基金在他职业生涯的关键阶段为这位研究人员提供了支持。复制启动涉及DNA和许多蛋白质之间的短暂相互作用，这对使用当前方法进行监测和测量具有挑战性。虽然在酵母中已经详细研究了复制起始的过程，但是关键蛋白在酵母和脊椎动物之间保守性很差，并且它们的确切功能仍然不清楚。本计画将利用主要研究者所开发之新颖单分子影像学方法，系统地剖析非洲爪蟾卵萃取物模型系统中的复制起始。这一努力将通过生物化学、基因组学和蛋白质组学方法得到补充。这项研究有可能通过以下方式推进该领域：（1）提供高等真核生物复制起始的定量视图，（2）鉴定和表征复制起始的未知调节因子，以及（3）该奖项反映了NSF的法定使命，并被认为值得通过使用基金会的学术价值和更广泛的影响审查标准。