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 复制染色体的组织和分离是生命系统的基础。结构 在生命的所有领域中，染色体的维持（SMC）复合体在这些过程中起着中心作用。 这些环状ATP酶具有共同的结构和亚基间的接触，与共同的 作用机制。最近在枯草芽孢杆菌和真核生物中的研究已经提供了令人信服的体内和体外研究结果。 SMC复合物利用ATP水解挤出DNA环的体外证据。然而，目前还不清楚如何 当染色体被许多蛋白质包被并受到蛋白质的作用时，SMC复合物挤出DNA环。 复制和转录机制。我的研究小组的目标是确定SMC复合物如何作用， 它们遇到复制体或其他凝聚素分子。我们还将描述SMC复合物是如何 当它们到达末端区域时从染色体中释放出来。最后，我们将探讨SMC在 组织和动力学的多部分细菌基因组，包含一个环形和线性 染色体总之，我们的工作有可能提供染色体折叠的一般原则 和致密化。 在此申请中，我要求行政补充，以支持本科生 在我的实验室里，林赛·韦女士，在2023年夏天在我的实验室里做一个研究项目。这个目标 该项目是为了支持女士的全职夏季研究经验的方式在细菌细胞生物学和分子 生物学路女士目前是IU的大三学生。她决心攻读博士学位。在生物医学科学中。完整的- 这个夏天的时间研究将更好地为她准备研究生院。韦女士的项目将调查 SMC复合物与来自细菌和古细菌的组蛋白样蛋白集合之间的相互作用， 包括来自E. coli的HMfA和HMfB。路女士将：1） 构建质粒和菌株以表达与mCherry融合的这些蛋白质; 2）使用 荧光显微镜; 3）使用Western印迹定量其表达; 4）监测 通过全基因组染色体构象捕获Hi-C分析来研究SMC-组蛋白相互作用。韦女士将 参加我们的每周实验室会议。她还将在印第安纳州微生物学大学展示一张海报 2023年10月的年度撤退。