Circadian gating of cell division by the cyanobacterial oscillator

描述（由申请人提供）：细胞分裂的时间与细胞生命中的其他周期事件协调。从细菌到藻类，再到哺乳动物的各种细胞类型，昼夜节律生物钟控制着一天中细胞分裂发生的时间。在任何系统中，这种时间限制或控制细胞分裂的机制和功能都知之甚少，而哺乳动物中的这种调节检查点是发育程序和癌症进展的重要因素。该项目的总体目标是了解昼夜节律和细胞分裂是如何以及为什么相互关联的。蓝藻长聚球菌细胞分裂的昼夜节律控制提供了一个优雅的系统，其中解决了探索时钟，细胞分裂机制和分离染色体的相互作用的问题。该项目旨在回答：细胞分裂的昼夜节律检查点的生物学作用是什么？连接生物钟和细胞分裂的机制有哪些组成部分？在细胞和昼夜周期中，时钟振荡器组件在哪里？它们是如何遗传的？此外，在倍性随昼夜节律波动的长叶蛇中，染色体的分裂是否与细胞分裂的门控有关？具体目标将：(1)定义昼夜节律钟调节细胞分裂的成分，并确定绕过细胞分裂大门的后果；(2)在昼夜节律和细胞分裂周期中识别时钟蛋白的细胞内定位和动态；(3)阐明生物钟、细胞质分裂和染色体分离之间的关系。现有的绕过昼夜节律细胞分裂检查点的突变体将被用于验证关于该检查点在保护昼夜节律精度、染色体完整性和/或基因表达的细胞间变异中的作用的假设。质谱法将确定与细胞相关的因素

DESCRIPTION (provided by applicant): The timing of cell division is coordinated with other cyclic events, of other periodicities, in the lives of cells. From bacteria, to algae, to diverse cll types in mammals, the circadian biological clock controls the time of day during which cell division can occur. The mechanism and function of this time restriction, or gating, of cell divisio is poorly understood in any system, and such regulatory checkpoints in mammals are important factors in developmental programs and the progression of cancer. The overall goal of this project is to understand how and why circadian rhythms and cell division are interlocked. The circadian control of cell division in the cyanobacterium Synechococcus elongatus provides an elegant system in which to address questions that probe the interactions of the clock, the cytokinesis machinery, and the segregating chromosomes. This project seeks to answer: What is the biological role of the circadian checkpoint of cytokinesis? What are the components of the machinery that connect the circadian clock to cell division? Where are the clock oscillator components during the cell and circadian cycles and how are they inherited? And, is the partitioning of chromosomes in S. elongatus, whose ploidy levels oscillate with circadian rhythmicity, related to the gating of cell division? The specific aims will: (1) Define the components, through which the circadian clock regulates cell division, and determine the consequences of bypassing the cell division gate; (2) Discern the intracellular localization and dynamics of clock proteins during the circadian and cell division cycles; and (3) Elucidate the relationships among the clock, cytokinesis, and chromosome segregation. Existing mutants that bypass the circadian cell division checkpoint will be used to test hypotheses that address the role of the gate in protecting circadian precision, chromosome integrity, and/or cell-to-cell variations in gene expression. Mass spectrometry will identify factors that associate with the cell division machinery during the gating checkpoint. Time-lapse imaging of cells trapped in microfluidics chambers will enable simultaneous monitoring of cell division and circadian rhythms to assess the consequences of gating in individual wild-type cells and in mutants that bypass the circadian gate. Super-resolution Structured Illumination imaging will provide sufficient sensitivity and resolution to track the localization and dynamics of circadian oscillato proteins and tagged chromosomes in wild-type and mutant cells throughout the circadian and cell division cycles. Sorting and imaging flow cytometry methods will be used to assess ploidy changes in wild-type and mutant strains. This project will reveal how and why a circadian clock controls cell division, a coupling of timing circuits that occurs in mammalian cells as well as in cyanobacteria, and will provide novel insight into how cells inherit a sense of time.