Mob1 Localization and Function During Mitosis

细胞分裂过程中染色体倍性的维持需要染色体的精确协调 分离和胞质分裂，使收缩环不组装之前，所有复制的姐妹 染色单体正确地附着到有丝分裂纺锤体上，并且满足纺锤体装配检查点。在 在芽殖和分裂酵母中，这一过程由称为有丝分裂退出的信号级联促进 网络/分隔起始网络协调有丝分裂退出和胞质分裂，基于纺锤体极 体，并在启动肌动蛋白环的收缩中发挥积极作用。迄今为止，很少有功能同源物具有 在动物细胞中进行了表征，但在末端组分（Mob 1和Dbf 2/Sid 2）的情况下， 在两个基因家族中都有扩展。虽然肿瘤抑制功能被归因于 哺乳动物和果蝇的直系同源物Mob 1和Dbf 2/Sid 2在G1/S，很少有人知道这些如何 分子参与动物细胞的有丝分裂和胞质分裂。在培养细胞中Mob 1亚型的初步研究 人类细胞表明，定位动力学反映了在酵母中观察到的，Mob 1在 有丝分裂早期的着丝粒和纺锤体极，以及胞质分裂期间的纺锤体中区和中体。 此外，我们已经确定，Mob 1和染色体乘客复合体是相互依赖的， 在有丝分裂早期相互定位。最后，我们确定了大肿瘤抑制因子2（Lats 2）作为一种肿瘤抑制因子。 Mob 1A特异性结合蛋白和Dbf 2/Sid 2激酶的可能功能同源物。利用这些初步 研究作为基础，该应用程序旨在联合收割机分子，生物化学和活细胞分析，以测试 假设Mob 1蛋白在维持染色体倍性的过程中起调节作用， 有丝分裂和间期。形成具体目标的实验路线将：1）确定 Mob 1定位于动粒的分子决定因素; 2）剖析Mob 1如何影响极光B 3）评估Mob 1参与Lats 2-p53应答， 细胞骨架破坏这些研究将为动物细胞中出现的基因家族提供新的见解。 作为细胞增殖的负调节剂（其功能丧失与肿瘤相关 形成），但对于完成酵母中的细胞分裂至关重要。预计这些研究将有助于我们 调和Mob 1如何能够参与这两个非常不同但绝对重要的细胞特征， 周期调节

Maintenance of chromosomal ploidy during cell division requires a precise coordination of chromosome segregation and cytokinesis such that the contractile ring does not assemble before all replicated sister chromatids are correctly attached to the mitotic spindle and the spindle assembly checkpoint is satisfied. In budding and fission yeast, this process is facilitated by a signaling cascade termed the Mitotic Exit Network/Septation Initiation Network that coordinates mitotic exit and cytokinesis, is based at the spindle pole bodies, and plays an active role in initiating constriction of the actin ring. To date, few functional homologues have been characterized in animal cells, but in the case of the terminal components (Mob1 and Dbf2/Sid2), there have been expansions in both gene families. And while tumor suppressor functions have been ascribed to the mammalian and Drosophila orthologs of Mob1 and Dbf2/Sid2 during G1/S, little is known about how these molecules participate in mitosis and cytokinesis in animal cells. Preliminary studies of Mob1 isoforms in cultured human cells indicate that the localization dynamics mirror that observed in yeast, with Mob1 enriched at the kinetochores and spindle poles early in mitosis, and the spindle midzone and midbody during cytokinesis. Moreover, we have determined that Mob1 and the chromosomal passenger complex are mutually dependent on each other kinetochore localization early in mitosis. Lastly, we identified Large Tumor Supressor 2 (Lats2) as a Mob1A-specific binding protein and possible functional homolog of the Dbf2/Sid2 kinase. Using these preliminary studies as a foundation, this application seeks to combine molecular, biochemical and live cell analyses to test the hypothesis that Mob1 proteins perform roles in regulating in maintaining chromosomal ploidy during both mitosis and interphase. The lines of experimentation that form the Specific Aims will: 1) Determine the molecular determinants of Mob1 localization to the kinetochore; 2) Dissect how Mob1 affects Aurora B function at the kinetochore; and 3) Assess the involvement of Mob1 in the Lats2-p53 response to cytoskeletal disruption. These studies will shed novel insights into a gene family that in animal cells appears to act as a negative regulator of cell proliferation (and whose loss of function is associated with tumor formation), yet is essential for completing cell division in yeast. It is anticipated that these studies will help us reconcile how Mob1 is capable of participating in both of these very different but absolutely critical features of cell cycle regulation.