How bacterial SMC complexes organize chromosomes

细菌 SMC 复合体如何组织染色体

• 批准号: 10809856
• 负责人: Xindan Wang
• 金额: $ 0.67万
• 依托单位: TRUSTEES OF INDIANA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-01 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10809856
• 关键词: ATP Hydrolysis; ATP phosphohydrolase; Administrative Supplement; Archaea; Bacillus subtilis; Bacteria; Bacterial Genome; Biological Assay; Cells; Cellular biology; Chimeric Proteins; Chromatin Loop; Chromosome Segregation; Chromosome Structures; Chromosomes; Collection; Complex; Doctor of Philosophy; Escherichia coli; Eukaryota; Fluorescence Microscopy; Funding; Genetic Transcription; Goals; Hi-C; Histones; In Vitro; Indiana; Life; Maintenance; Microbiology; Molecular Biology; Monitor; Organism; Plasmids; Play; Process; Proteins; Research; Research Project Grants; Role; Science; Shapes; Structure; Universities; Visualization; Western Blotting; Work; chromosome conformation capture; chromosome replication; condensin; experience; genome integrity; genome-wide; graduate school preparation; in vivo; meetings; posters; segregation; summer research; undergraduate research experience; undergraduate student

Project Summary The organization and segregation of replicated chromosomes are fundamental to living systems. Structural maintenance of chromosomes (SMC) complexes play central roles in these processes in all domains of life. These ring-shaped ATPases share common structures and inter-subunit contacts, consistent with a common mechanism of action. Recent studies in Bacillus subtilis and eukaryotes have provided compelling in vivo and in vitro evidence that SMC complexes utilize ATP hydrolysis to extrude DNA loops. However, it is unclear how SMC complexes extrude DNA loops when the chromosome is coated by numerous proteins and acted upon by replication and transcription machineries. My research group aims to determine how SMC complexes act when they encounter the replisome or other condensin molecules. We will also characterize how SMC complexes are released from the chromosome when they reach the terminus region. Finally, we will explore SMC’s role in the organization and dynamics of a multipartite bacterial genome that contains both a circular and a linear chromosome. Taken together, our work has the potential to provide the general principles of chromosome folding and compaction in all organisms. In this proposed application, I am requesting an administrative supplement to support an undergraduate in my lab, Ms. Lindsey Way, to do a research project in my lab during the summer of 2023. The goal of this project is to support Ms. Way for full-time summer research experience in bacterial cell biology and molecular biology. Ms. Way is currently a Junior at IU. She is determined to pursue a Ph.D. in biomedical sciences. A full- time research in this summer will better prepare her for graduate school. Ms. Way’s project will investigate the interactions between SMC complexes and a collection of histone-like proteins from bacteria and archaea, including HupA and H-NS from E. coli, and HMfA and HMfB from Methanothermus fervidus. Ms. Way will: 1) build plasmids and strains to express these proteins fused to mCherry; 2) visualize these fusion proteins using fluorescence microscopy; 3) quantify their expression using Western blotting; 4) monitor the consequences of the SMC-histone interactions by genome-wide chromosome conformation capture Hi-C assays. Ms. Way will present in our weekly lab meetings. She will also present a poster in the Indiana University Microbiology Annual Retreat in October 2023.

项目摘要 复制的染色体的组织和隔离是生物系统的基础。结构 维持染色体（SMC）复合物在生活的所有领域的这些过程中都起着核心作用。 这些环形ATPases共享共同的结构和育间接触，与共同 作用机理。最近对枯草芽孢杆菌和真核生物的研究为体内和 SMC复合物利用ATP水解挤出DNA环的体外证据。但是，目前尚不清楚如何 当染色体被多种蛋白覆盖并作用于 复制和转录机。我的研究小组旨在确定SMC复合物的行为 它们遇到复制体或其他冷凝蛋白分子。我们还将表征SMC复合体 当它们到达末端区域时，它们从染色体释放。最后，我们将探讨SMC在 多部分细菌基因组的组织和动力学，既包含圆形和线性 染色体。综上所述，我们的工作有可能提供染色体折叠的一般原理 和所有生物体的压实。 在此提议的申请中，我要求提供管理补充剂以支持本科 在我的实验室，Lindsey Way女士中，在2023年夏天在我的实验室中进行研究项目。 项目是为了支持细菌细胞生物学和分子的全日制夏季研究经验的道路 生物学。 Way女士目前是IU的大三学生。她决心攻读博士学位。在生物医学科学中。一个完整的 这个夏天的时间研究将更好地为研究生院做好准备。 Way女士的项目将调查 SMC复合物与细菌和古细菌的组蛋白样蛋白的相互作用， 包括来自大肠杆菌的HUPA和H-NS，以及来自甲壳虫肠的HMFA和HMFB。 Way女士Will：1） 建立质粒和菌株以表达这些蛋白质融合到麦克里； 2）使用 荧光显微镜； 3）使用蛋白质印迹量化其表达； 4）监视后果 全基因组染色体会议捕获HI-C测定法的SMC-Histone相互作用。女士Will 在我们的每周实验室会议中出席。她还将在印第安纳大学微生物学上介绍一张海报 2023年10月的年度撤退。