Novel Regulatory Factors of the Spindle Assembly Checkpoint

 描述（由申请人提供）：人类细胞分裂是一组高度协调的事件，确保遗传物质（染色体）从一个母细胞正确传递到两个新形成的细胞。细胞分裂期间的染色体错误分离可导致非整倍性（染色体数目异常），这是大多数癌症的标志，并且已被提出促进肿瘤发生。在中期到后期的过渡期确保正确的姐妹染色单体分离的关键是多组分纺锤体组装检查点（SAC），当未连接的动粒或非生产性（单端，syntelic和merotelic）附件被感知时，它被激活，并在中期阻滞细胞，以便有时间纠正这些缺陷，并在细胞分裂前产生适当的微管-动粒附件。有趣的是，一个功能性的SAC在有效性方面发挥着作用， 化疗药物如抗有丝分裂剂（抑制有丝分裂的药物），其破坏有丝分裂纺锤体，激活SAC，将细胞阻滞在前中期并触发凋亡性细胞死亡。由于了解SAC对于了解肿瘤发生和肿瘤细胞对抗有丝分裂药物的反应至关重要，因此它已成为一个有吸引力的研究领域。尽管过去40年的研究已经阐明了SAC，但我们还远未阐明参与这一复杂途径的调节因子的全部补充，也远未了解这一途径的错误调节如何导致肿瘤发生和对化疗药物如抗有丝分裂剂的耐药性。为了解决这些问题，我们最近进行了一个高通量的小干扰RNA（siRNA）的SAC的新监管机构的屏幕。该方法产生了两种新的细胞周期蛋白依赖性激酶（Cdk 14和Cdk 15）和两种双特异性磷酸酶（DUSP 7和DUSP 12）。这些新因子的失活导致SAC旁路在抗有丝分裂药物如紫杉醇的存在下，其通常激活SAC并诱导细胞死亡。据我们所知，这些因素以前没有与SAC功能相关，并且通常具有有限的遗传和分子表征。因此，了解这些因素如何调节SAC对于理解SAC和更广泛的细胞分裂很重要。我们推测这些因子通过其磷酸化和去磷酸化活性控制SAC。因此，我们将通过分析这些功能来验证这一假设。 在未受干扰的细胞分裂过程中以及在产生纺锤体损伤并激活纺锤体组装检查点的抗有丝分裂剂存在下，蛋白质被破坏。我们希望我们的研究能够识别和表征调控SAC的新因子和途径，我们将能够绘制它们在时间和空间中的作用，并且我们对理解细胞分裂机制的贡献将影响未来的癌症研究和癌症患者的生活质量和生存率。

 DESCRIPTION (provided by applicant): Human cell division is a highly coordinated set of events that ensures the proper transmission of genetic material (chromosomes) from one mother cell to two newly formed cells. Chromosome missegregation during cell division can lead to aneuploidy (an aberrant chromosomal number), which is a hallmark of most cancers and has been proposed to promote tumorigenesis. Critical to ensuring proper sister chromatid separation at the metaphase to anaphase transition is the multi-component spindle assembly checkpoint (SAC), which is activated when unattached kinetochores or nonproductive (monotelic, syntelic, and merotelic) attachments are sensed and functions to arrest cells in metaphase to give time to correct these deficiencies and generate proper microtubule-kinetochore attachments before proceeding with cell division. Interestingly, a functional SAC plays a role in the effectiveness of chemotherapeutic drugs like antimitotics (drugs that inhibit mitosis), which damage the mitotic spindle, activate the SAC, arrest cells in prometaphase and trigger apoptotic cell death. Because understanding the SAC is critical to understanding tumorigenesis and the response of tumor cells to antimitotic drugs, it has become an attractive area of research. Although the last 40 years of research has shed light on the SAC, we are far from elucidating the full complement of regulatory factors involved in this complex pathway and from understanding how misregulation of this pathway can lead to tumorigenesis and resistance to chemotherapeutic drugs like antimitotics. To address these issues, we recently performed a high-throughput small interfering RNA (siRNA) screen for novel regulators of the SAC. This approach yielded two novel cyclin dependent kinases (Cdk14 and Cdk15) and two dual specificity phosphatases (DUSP7 and DUSP12). Inactivation of these novel factors leads to SAC bypass in the presence of antimitotic drugs like Taxol, which normally activate the SAC and induce cell death. To our knowledge, these factors have not been previously linked to SAC functioning and have limited genetic and molecular characterization in general. Thus understanding how these factors regulate the SAC is important to understanding the SAC and more broadly cell division. We hypothesize that these factors are controlling the SAC through their phosphorylation and dephosphorylation activities. Thus, we will test this hypothesis by analyzing the function of these proteins during unperturbed cell divisions and in the presence of antimitotics that generate spindle damage and activate the spindle assembly checkpoint. We expect that our studies will identify and characterize new factors and pathways that regulate the SAC, that we will be able to map their roles in time and space, and that our contributions to understanding the mechanisms of cell division will impact future cancer studies and cancer patient quality of life and survival.