Roles of chromosomal factors in chromosome segregation

染色体因素在染色体分离中的作用

• 批准号: 8708102
• 负责人: Hironori Funabiki
• 金额: $ 34.75万
• 依托单位: ROCKEFELLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-08-01 至 2015-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8708102
• 关键词: Address; Affect; Anaphase; Aneuploidy; Apoptosis; Apoptotic; BIR Domain; Binding; Biochemical; Biological; CDC2 Protein Kinase; Cell Proliferation; Cell division; Cells; Centromere; Chromatin; Chromosome Segregation; Chromosomes; Coiled-Coil Domain; Complex; Congenital Abnormality; Coupled; Data; Detection; Development; Embryo; Embryonic Development; Encapsulated; Ensure; Failure; Fertilization; Growth; Haspin; Histone H3; Kinetochores; Ligands; Link; Malignant Neoplasms; Maps; Mass Spectrum Analysis; Mediating; Meiosis; Metaphase; Methods; Microtubules; Mitosis; Mitotic; Molecular; Monitor; Mutation; N-terminal; Normal Cell; Nuclear; Nuclear Envelope; Nuclear Magnetic Resonance; Outcome; Pathway interactions; Phosphorylation; Phosphotransferases; Play; Prevention; Process; Protein Kinase; Proteins; Publishing; Regulation; Role; Signal Pathway; Signal Transduction; Sister; Spatial Distribution; Specificity; Staging; Structure; Testing; Therapeutic procedure; Threonine; Work; Xenopus; Xenopus laevis; aurora B kinase; base; cancer cell; cell type; chemical reaction; chromosome movement; egg; improved; inner centromere protein; insight; killings; mutant; neutralizing antibody; novel; pluripotency; response; signal processing; spatiotemporal; survivin; tumor

DESCRIPTION (provided by applicant): Chromosomes actively generate a variety of essential signals that promote spindle assembly and chromosome movement during mitosis and meiosis. Failure to properly complete these processes leads to aneuploidy, which is frequently associated with cancer and birth defects. In this proposal, we will focus on a critical regulator of these processes, the chromosomal passenger complex (CPC), consisting of the kinase Aurora B, and its regulatory subunits, INCENP, Dasra and Survivin. The CPC localizes to chromatin and is enriched at centromeres. Chromatin binding is directly coupled to activation of the kinase activity of Aurora B. In Xenopus laevis egg extracts, this activation mechanism is critical for spindle assembly. At centromeres, the CPC mediates spindle assembly checkpoint (SAC) signaling to delay anaphase onset in response to erroneous kinetochore attachments. To reveal the spatiotemporal mechanism that controls the Aurora B pathway, we will: 1) Address how the CPC controls the SAC. We have found that the putative coiled-coil (CC) domain of INCENP is critical for the SAC. To test if the CC domain monitors physical kinetochore changes upon microtubule attachment and converts them into Aurora B signaling, the CC domain will be systematically modulated, and changes in Aurora B-dependent phosphorylation within kinetochores will be monitored. 2) Reveal a mechanism by which a network of protein kinases controls M phase-specific activation of Aurora B by chromatin. Based on our preliminary data, we hypothesize that the histone H3 threonine 3 (H3T3) kinase Haspin is activated by the mitotic kinases Cdk1 and Plx1 to mediate the temporal control of Aurora B activation by chromatin. The precise mechanism by which these mitotic kinases activate Haspin will be analyzed by biochemical methods combined with advanced mass spectrometry. 3) Provide the structural basis for the CPC- chromosome interaction. Localization of the CPC to chromosomes is mediated by Survivin, which directly binds to histone H3 that has been phosphorylated at threonine 3 (H3T3ph). To determine how this phosphorylation regulates binding specificity, crystal structures of the Survivin-H3T3ph complex will be determined. 4) Investigate another chromatin-activated protein that was recently identified in my lab, Vespera, which appears to oppose the Aurora B pathway, inhibit microtubule assembly, and promote nuclear re- formation. Since Vespera is exclusively expressed in pluripotent cells (including eggs), it is possible that its presence is linked to the requirement for the CPC in spindle assembly in these cell types. Therefore we will investigate specific roles for Vespera during these developmental stages. The outcomes of this proposed project will provide molecular explanations for how the chromosome-associated activities of the CPC and Vespera coordinate mitosis, and will help us understand how their misregulation affects the growth of normal and cancer cells.

描述（由申请人提供）：染色体积极产生各种基本信号，这些信号在有丝分裂和减数分裂过程中促进主轴组件和染色体运动。无法正确完成这些过程会导致非整倍性，这通常与癌症和先天缺陷有关。在此提案中，我们将重点关注这些过程的关键调节剂，即由激酶Aurora B及其调节亚基，Incenp，Dasra和Survivin组成的染色体乘客复合物（CPC）。 CPC定位于染色质，并在丝粒中富集。染色质结合直接与AuroraB的激酶活性的激活直接耦合。在Xenopus laevis卵提取物中，这种激活机制对于纺锤体组装至关重要。在CentRomeres，CPC介导主轴组件检查点（SAC）信号传导以响应错误的动力学附件延迟后期发作。为了揭示控制Aurora B途径的时空机制，我们将：1）解决CPC如何控制SAC。我们发现，Incenp的推定盘绕圈（CC）域对于SAC至关重要。为了测试CC结构域在微管附着时是否监视物理动力学变化并将其转换为Aurora b信号传导，将系统调节CC域，并且将监测动力学内化学中Aurora B依赖性磷酸化的变化。 2）揭示了一种机制，通过这种机制，蛋白激酶网络通过染色质控制Aurora B的M相特异性激活。基于我们的初步数据，我们假设组蛋白H3苏氨酸3（H3T3）激酶HASPIN被有丝分裂激酶CDK1和PLX1激活，以介导染色质激活Aurora B激活的时间控制。这些有丝分裂激酶激活HASPIN的确切机制将通过生化方法与晚期质谱法相结合。 3）为CPC-染色体相互作用提供结构基础。 CPC对染色体的定位是由Survivin介导的，Survivin直接与在苏氨酸3（H3T3PH）磷酸化的组蛋白H3结合。为了确定这种磷酸化如何调节结合特异性，将确定Survivin-H3T3PH复合物的晶体结构。 4）研究了最近在我的实验室Vespera中鉴定出的另一种染色质激活的蛋白质，该蛋白似乎反对Aurora B途径，抑制微管组装并促进核能。由于Vespera在多能细胞（包括卵）中仅表达，因此其存在可能与这些细胞类型中纺锤体组装中CPC的需求有关。因此，我们将在这些发展阶段研究Vespera的特定作用。该提出的项目的结果将为CPC和Vespera协调有丝质的染色体相关活性提供分子解释，并将帮助我们了解它们的正常状态如何影响正常和癌细胞的生长。