Topoisomerases and Chromosome Segregation

拓扑异构酶和染色体分离

• 批准号: 7671615
• 负责人: KENNETH J MARIANS
• 金额: $ 48.29万
• 依托单位: SLOAN-KETTERING INST CAN RESEARCH
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-07-01 至 2013-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7671615
• 关键词: Ablation; Accounting; Address; Alleles; Antibiotics; Antitoxins; Bacteria; Bacterial Chromosomes; Biochemical; Biological; C-terminal; Cell Cycle; Cell division; Cells; Cellular Morphology; Chromosome Segregation; Chromosomes; Coupled; Coupling; Cytoskeletal Proteins; Cytoskeleton; DNA; DNA Binding; DNA Topoisomerase IV; DNA biosynthesis; Defect; Dependency; Elements; Ensure; Entropy; Enzymes; Escherichia coli; Eukaryota; Event; Flow Cytometry; Genes; Genetic; Genetic Screening; Genetic Transcription; Goals; Grant; In Vitro; Integral Membrane Protein; Investigation; Mediating; Mediator of activation protein; Membrane; Molecular; Molecular Genetics; Molecular Motors; Morphology; Motor; Mutation; Nature; Participant; Pathway interactions; Physical condensation; Process; Property; Proteins; Public Health; Reaction; Regulation; Resolution; Role; SS DNA BP; Sister; Sorting - Cell Movement; Spatial Distribution; Staging; Techniques; Temperature; Topoisomerase; Topoisomerase III; Toxin; Variant; Yeasts; antimicrobial; antimicrobial drug; bacterial resistance; base; chemotherapy; condensin; daughter cell; dimer; in vivo; mutant; overexpression; segregation; transmission process; yeast two hybrid system

DESCRIPTION (provided by applicant): Accurate chromosome segregation is crucial to ensure that each daughter cell receives a complete copy of the genetic information. Molecular mechanisms that drive these processes are well understood in eukaryotes, however, we know little about these events in bacteria. Recent advances in the application of cell biological techniques have revealed that the spatial distribution of the bacterial chromosome is highly ordered and that chromosome segregation is likely to be a progressive process. The force for DNA segregation has been ascribed to replication itself, transcription, transertion (the coupling of transcription and co-translational insertion of proteins into the membrane], and entropy. And the role of the recently discovered MreB cytoskeleton remains unclear. The long term goal of this grant is to understand the events necessary for proper segregation of the newly duplicated Escherichia coli sister chromosomes to a new daughter cell. Our focus in these investigations is topoisomerase IV (Topo IV), which is responsible for unlinking the catenated sister chromosomes, and its interactions with other proteins involved in chromosome dynamics and cell division. In the previous grant period we have: 1) shown that Topo IV activity is regulated by the oligomeric state of the cytoskeletal element MreB, monomeric MreB inhibits whereas filamentous MreB stimulates, possibly accounting for the temporal regulation of Topo IV activity in the cell; 2) discovered and characterized an interaction between the ParC subunit of Topo IV and MukB, the bacterial condensin, that stimulates Topo IV activity; 3) demonstrated that stimulation of Topo IV by FtsK, the molecular motor required for final sorting of the chromosomes, does not require FtsK DNA translocation; 4) identified a nucleoid associated protein, YejK, that interacts with Topo IV and demonstrated that ??yejK cells have a cell cycle defect; 5) demonstrated biochemically that RecQ and topoisomerase III (Topo III) can resolve convergent replication forks, a reaction that may be important at the terminal stages of DNA replication; and 6) discovered that regulation of cell division is coupled to the condensation state of the nucleoid, possibly via a checkpoint that, when engaged, inhibits Min protein oscillation, required for proper placement of the division septum. We will proceed to use a combination of biochemical, cell biologic, and molecular genetic approaches to answer the following questions: What is the role of Topo IV in chromosome dynamics and how is this role modulated by the interactions between Topo IV and MukB and Topo IV and FtsK? How is Topo IV activity regulated in the cell and what is the role of the Topo IV-MreB interaction in this process? What is the nature of the coupling between cell division and the condensation state of the nucleoid? And, do RecQ and Topo III support an alternate pathway of sister chromosome decatenation in vivo? PUBLIC HEALTH REVELANCE: Resistance of bacteria to treatment with antibiotic and anti-microbial drugs is a persistent public health problem that is increasingly of concern. This proposal investigates emerging new paradigms in the bacterium Escherichia coli that are involved in the accurate transmission of the genetic information to the daughter cells. We anticipate that as we understand more about these pathways and more of the participants are revealed, potential new targets for antimicrobial chemotherapy will be presented.

描述（由申请人提供）：准确的染色体分离对于确保每个子细胞都会收到遗传信息的完整副本至关重要。驱动这些过程的分子机制在真核生物中被充分理解，但是，我们对细菌中的这些事件知之甚少。细胞生物技术应用的最新进展表明，细菌染色体的空间分布高度有序，并且染色体分离可能是一个渐进的过程。 DNA分离的力已归因于复制本身，转录，转置（转录和将蛋白质的共插入蛋白偶联到膜中]和熵。以及最近发现的MREB Cytoskeleton的作用，这是不清楚的长期竞赛的长期竞赛。我们在这些研究中的重点是topoisomerase IV（topo iv），该染色体是为链接的，它与染色体动力学和细胞分裂相互作用的蛋白质与其他蛋白质相互作用。抑制丝状MREB刺激，可能会考虑细胞中托普IV活性的时间调节。 2）发现并表征了刺激Topo IV活性的细菌冷凝蛋白的TOPO IV和MUKB的PARC亚基之间的相互作用； 3）证明，染色体最终排序所需的分子运动所需的ftsk刺激TOPO IV不需要FTSK DNA易位。 4）鉴定出与Topo IV相互作用的核苷蛋白Yejk，并证明了Yejk细胞具有细胞周期缺陷； 5）从生物化学上证明了RECQ和TOPOISomerase III（TOPO III）可以解决收敛复制叉，这种反应在DNA复制的末端可能很重要。 6）发现细胞分裂的调节与核苷的冷凝状态耦合，这可能是通过检查点，即在接合时抑制最小蛋白质振荡，以适当放置分裂的隔膜。我们将开始使用生化，细胞生物学和分子遗传方法的结合来回答以下问题：topo iv在染色体动力学中的作用是什么，以及这种作用如何受到topo IV和MUKB和MUKB和MUKB和MUKB和TOPO IV和TOPO IV和TOPO IV和FTSK之间的相互作用？ TOPO IV活性如何在细胞中调节？TOPO IV-MREB相互作用在此过程中的作用是什么？细胞分裂和核苷的冷凝态之间耦合的性质是什么？而且，RECQ和TOPO III是否支持体内姐妹染色体衰减的替代途径？ 公共卫生启示：细菌对抗生素和抗菌药物治疗的抵抗力是越来越令人关注的持续公共卫生问题。该提案调查了大肠菌中新兴的新范式，这些范式参与了将遗传信息准确地传播给子细胞的。我们预计，随着我们对这些途径的更多了解，并且更多的参与者被揭示了抗菌剂的潜在新目标 将提出化学疗法。