Cell Cycle Regluation in Caulobacter crescentus

描述（由申请人提供）：适当的细胞周期的时空调节对所有生物的生存和增殖至关重要。为了确保稳定有序的细胞周期进程，细胞使用复杂的遗传回路。这种电路的结构、操作和设计原理仍然知之甚少。月牙根杆菌为研究遗传回路和阐明细胞周期调控的基本分子机制提供了一个实验可处理的系统。在月牙草中，细胞周期是由主调控因子CtrA活性的周期性上升和下降驱动的。活性CtrA直接调节近100个基因的表达，其中许多基因协调细胞分裂。CtrA也直接结合并沉默复制的起始点。因此，细胞周期的进展需要CtrA活性的振荡——它必须从细胞中清除以允许DNA复制，但必须积累以驱动细胞分裂。控制CtrA的遗传电路的轮廓已经确定，但还不能完全解释CtrA活性的动态。必须存在额外的组成部分和监管水平。该项目的目标是确定控制CtrA活性的完整分子电路，从而确保细胞周期的有序进展。为此，我们将：(i)绘制调节CtrA磷酸化和去磷酸化的途径，（ii）阐明细胞如何控制CckA （CtrA的主要磷供体）的亚细胞定位和活性，（iii）研究多个转录反馈回路如何协作以精确控制CtrA的诱导动力学，以及（iv）识别和表征调节CtrA活性的其他因素。这些研究将有助于揭示细胞如何使用多种调节模式来成功地驾驭它们的细胞周期。此外，这项工作将有助于揭示遗传电路的一般设计和工作原理，这是整个生物学调节过程的基础。最后，更好地了解细菌如何调节细胞周期可能有助于指导新抗生素的开发，这是一个与公共卫生日益相关的问题。

DESCRIPTION (provided by applicant): Proper spatial and temporal regulation of the cell cycle is crucial to the survival and proliferation of all organisms. To ensure stable, orderly cell cycle progression, cells employ complex genetic circuits. The architecture, operation, and design principles of such circuits remain only poorly understood. The bacterium Caulobacter crescentus provides an experimentally tractable system for studying genetic circuits and for elucidating fundamental molecular mechanisms of cell cycle regulation. In C. crescentus the cell cycle is driven by the periodic rise and fall in activity of a master regulator, CtrA. Active CtrA directly regulates the expression of nearly 100 genes, many of which coordinate cell division. CtrA also directly binds to and silences the origin of replication. Cell cycle progression therefore requires oscillations in CtrA activity - it must be cleared from the cell to permit DNA replication, but must accumulate to drive cell division. The outline of a genetic circuit controlling CtrA is in place, but does not yet fully account for the dynamics of CtrA activity. Additional components and levels of regulation must exist. The goal of this project is to define the complete molecular circuitry that controls CtrA activity and hence ensures orderly progression through the cell cycle. To this end, we will: (i) map the pathways that regulate phosphorylation and dephosphorylation of CtrA, (ii) elucidate how cells control the sub-cellular localization and activity of CckA, the primary phosphodonor for CtrA (iii) examine how multiple transcriptional feedback loops collaborate to precisely control the induction dynamics of CtrA, and (iv) identify and characterize additional factors that regulate CtrA activity. These studies will help to unveil how cells use multiple modes of regulation to successfully navigate their cell cycle. In addition, this work will help to reveal the general design and operating principles of genetic circuits, which underlie regulatory processes throughout biology. Finally, a better understanding of how bacteria regulate the cell cycle may help guide the development of new antibiotics, a problem of increasing public-health relevance.