Regulation of cell division by mitotic kinases

描述(申请人提供)：细胞分裂的调节对于维持基因组的完整性是必要的。调控缺陷会导致染色体不平等分离和非整倍体，这与人类有丝分裂中的癌症、减数分裂中的妊娠丢失和发育缺陷密切相关。许多复杂的细胞分裂动力学是由几个关键的调节激酶控制的。其中的一种，极光B，定位于着丝粒直到着丝粒的后期，并调节染色体与有丝分裂纺锤体的附着。在后期，Aurora B重新分布到纺锤体中部，控制后期微管动力学和细胞质分裂。激酶定位和与其他调节蛋白相互作用的变化表明，磷酸化模式不同，这将允许单个激酶在不同的时间和地点调节多个过程。这项建议侧重于研究活细胞中具有高时间和空间分辨率的磷酸化动力学，以测试如何控制这些动力学的模型。极光B的一个关键作用是确保所有染色体以正确的构型附着在纺锤体微管上，以便染色体在后期准确分离。必须产生信号来区分正确和不正确的连接，以便纠正错误。第一个具体目标是测试单个着丝粒上的差异极光B信号模型。纺锤体微管施加的力在着丝粒上产生张力，这可能通过将底物与集中在着丝粒内侧的激酶物理分离来调节着丝粒底物的磷酸化。第二个具体目标是研究Aurora B和着丝粒上的其他有丝分裂酶之间的相互作用。有证据表明Aurora B、BubR1、Plk1和Chk1之间存在相互作用。通过使用小分子抑制剂和RNAi，将活细胞中磷酸化动力学的定量测量与激酶抑制相结合，将测试一个模型，在该模型中，多个激酶在着丝粒处作用于信号网络。Aurora B的功能和定位在后期发生了显著变化，这表明复杂的空间和时间磷酸化模式可能允许单一的a激酶调节多个细胞过程。第三个具体目标将确定Aurora B和相反的磷酸酶如何在后期位置特异性磷酸化动力学的切换中做出贡献。这些研究将有助于理解(1)调节细胞分裂的基本机制和(2)正在进行癌症治疗临床试验的Aurora B抑制剂的效果。 与公共卫生相关：对细胞分裂的适当调控确保子代细胞继承正确的遗传物质。分裂过程中的错误会导致细胞出现遗传异常，这些异常与人类癌症、妊娠丢失和发育缺陷密切相关。这项提议的目标是了解一种关键调节蛋白在细胞分裂中的功能，这是癌症治疗的一个有希望的靶点。

DESCRIPTION (provided by applicant): Regulation of cell division is necessary to maintain genome integrity. Defective regulation leads to unequal chromosome segregation and aneuploidy, which is strongly associated with human cancer in mitotic divisions and pregnancy loss and developmental defects in meiosis. Much of the complex dynamics of cell division is controlled by a few key regulatory kinases. One of these kinases, Aurora B, localizes to the centromere until anaphase and regulates the attachments of chromosomes to the mitotic spindle. At anaphase onset Aurora B redistributes to the spindle midzone and controls anaphase microtubule dynamics and cytokinesis. Changes in kinase localization and interactions with other regulatory proteins suggest varying patterns of phosphorylation that would allow a single kinase to regulate multiple processes at different times and places. This proposal focuses on examining phosphorylation dynamics with high temporal and spatial resolution in living cells to test models for how these dynamics are controlled. A key role of Aurora B is to ensure that all chromosomes attach to spindle microtubule in the correct configuration, so that chromosomes segregate accurately in anaphase. Signals must be generated to distinguish correct and incorrect attachment, so that errors can be corrected. The first specific aim will test a model for differential Aurora B signaling at individual centromeres. Forces exerted by spindle microtubules create tension across the centromere, which may regulate phosphorylation of a kinetochore substrate by physically separating the substrate from the kinase, which is concentrated at the inner centromere. The second specific aim will examine the interplay between Aurora B and other mitotic kinases at the centromere. There is evidence for interactions between Aurora B, BubR1, Plk1, and Chk1. By combining quantitative measurements of phosphorylation dynamics in living cells with kinase inhibition, using small molecule inhibitors and RNAi, a model in which multiple kinases act in a signaling network at the centromere will be tested. Aurora B function and localization change dramatically in anaphase, suggesting that complex spatial and temporal phosphorylation patterns may allow a single a kinase to regulate a multiple cellular process. The third specific aim will determine how both Aurora B and opposing phosphatases contribute to a switch in site- specific phosphorylation dynamics in anaphase. These studies will contribute to an understanding of (1) basic mechanisms regulating cell division and (2) the effects of Aurora B inhibitors, which are in clinical trials for cancer therapy. Public Health Relevance: Proper regulation of cell division ensures that daughter cells inherit the correct genetic material. Errors during division lead to cells with genetic abnormalities that are strongly associated with human cancer, pregnancy loss, and developmental defects. The goal of this proposal is to understand the function of a key regulatory protein, which is a promising target for cancer therapy, in cell division.