Identifying molecular mechanisms that suppress centriole amplification.

识别抑制中心粒扩增的分子机制。

• 批准号: 9267488
• 负责人: Gregory Charles Rogers
• 金额: $ 28.52万
• 依托单位: UNIVERSITY OF ARIZONA
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-05-01 至 2020-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9267488
• 关键词: Aneuploidy; Binding; Biochemical; Biological Assay; Biological Models; Cell Cycle; Cell Cycle Progression; Cell Cycle Regulation; Cell division; Cells; Centrioles; Centrosome; Chromosomal Instability; Chromosome Segregation; Chromosomes; Cilia; Complement; Complex; Congenital Abnormality; Coupled; DNA biosynthesis; Dangerousness; Data; Daughter; Defect; Disease; Drosophila genus; Drosophila polo protein; Elements; Ensure; Etiology; Event; Future; Genome; Goals; Growth; In Vitro; Inherited; Island; Knowledge; Laboratories; Licensing; Link; Location; Mediating; Meiosis; Microcephaly; Microtubules; Mission; Mitosis; Mitotic; Modification; Molecular; Mothers; National Institute of General Medical Sciences; Normal Cell; Oncogenic; Organelles; PLK1 gene; Pathway interactions; Pattern; Phosphorylation; Population; Process; Production; Proteins; Public Health; Regulation; Regulatory Pathway; Research; Resistance; Role; S Phase; Scaffolding Protein; Seckel syndrome; Series; Site; Spontaneous abortion; Spottings; Surface; System; Testing; Ubiquitin; Work; anaphase-promoting complex; cancer cell; ciliopathy; daughter cell; driving force; experimental study; in vivo; insight; mutant; novel; overexpression; prevent; programs; protein degradation; public health relevance; stem cell division; tumorigenesis; ubiquitin ligase

 DESCRIPTION (provided by applicant): Centrioles are the duplicating elements of centrosomes, organelles which nucleate microtubule growth to build mitotic/meiotic spindles and cilia. During each normal cell division, centrioles duplicate once and once only. Like DNA replication, centriole duplication is a semi-conservative process tightly coupled to the cell cycle and occurs only during S-phase. Centriole number must be closely regulated because a deficiency or an excess of centrioles may interfere with correct chromosome segregation and asymmetric stem cell division. The presence of too few centrioles contributes to various ciliopathies, including microcephaly, and Seckel syndrome. In contrast, too many centrioles directly promotes chromosomal instability (CIN) a driving force for aneuploidy that induces miscarriage, birth defects, and tumorigenesis. Furthermore, the presence of too many centrioles is frequently observed in cancer cells. Normally, each `mother' centriole assembles only a single `daughter', but centrioles have the capacity to assemble multiple daughters simultaneously - an aneuploidy- generating condition called centriole amplification. Normally, only a single, restricte site on the mother centriole is used to spawn a daughter. However, the mechanism limiting assembly to only this site is unknown. Experiments in this proposal test a hypothesis that explains why centriole duplication is normally restricted to a single event per mother centriole. Specifically, we link three conserved centriolar proteins, Polo-like kinase 4 (Plk4), its binding-partner and scaffolding protein Asterless (Asl), and the centriole assembly protein Anastral Spindle 2 (Ana2) into a single regulatory pathway. Overexpression of any of these proteins induces assembly of multiple daughter centrioles. How the cell cycle control program regulates these key proteins to limit daughter centriole assembly to only a single event is unknown. Here, we hypothesize that cell cycle regulators, including the Anaphase Promoting Complex (APC) ubiquitin-ligase, control the destruction of a Plk4/Asl/Ana2 complex to alter their pattern/activit on mother centrioles and generate a single site for daughter assembly. Our preliminary data indicate that 1) the Plk4 pattern on centrioles changes during mitotic progression: first coating the entire mother centriole but then being pruned to a single asymmetric spot by mitotic exit; 2) Plk4 and Ana2 are APC targets; 3) phosphorylation protects subpopulations of these proteins from degradation; and 4) Asl is a Plk4 substrate that stabilizes mitotic Plk4. In this proposal, we will (1) determine if APC regulation of Plk4 localization on centrioles defines the site of daughte centriole assembly, (2) determine the functional interaction between Plk4 and Asl which assists in defining a single site of centriole assembly, and (3) determine if Ana2 regulates Plk4 activation and whether Ana2 is controlled by APC. These studies will provide mechanistic insight into a molecular process that governs the fidelity of centriole duplication. Understanding the process at the molecular level will guide future studies to explore the etiology of centriole amplification during tumorigenesis and other centriole-related diseases.

 描述（由申请人提供）：中心粒是中心体的复制元件，是使微管生长成核以构建有丝分裂/减数分裂纺锤体和纤毛的细胞器。在每次正常细胞分裂期间，中心粒复制一次且仅复制一次。与DNA复制一样，中心粒复制是一个与细胞周期紧密相关的半保守过程 并且仅在S阶段期间发生。中心粒数量必须受到严格的调控，因为中心粒的缺乏或过量可能会干扰正确的染色体分离和不对称的干细胞分裂。中心粒过少会导致各种纤毛病变，包括小头畸形和塞克尔综合征。相反，太多的中心粒直接促进染色体不稳定性（CIN），这是诱导流产、出生缺陷和肿瘤发生的非整倍体的驱动力。此外，在癌细胞中经常观察到太多中心粒的存在。正常情况下，每个“母亲”中心粒只组装一个“女儿”，但中心粒有能力同时组装多个女儿-一种称为中心粒扩增的非整倍体产生条件。通常情况下，只有一个单一的，限制性的网站上的母亲中心粒是用来产卵的女儿。然而，机制限制大会只有这个网站是未知的。在这个提议中的实验测试一个假设，解释了为什么中心粒复制通常限于一个单一的事件，每个母亲中心粒。具体而言，我们将三个保守的中心粒蛋白，Polo样激酶4（Plk 4），其结合伙伴和支架蛋白Asterless（Asl），和中心粒组装蛋白Anastral Spindle 2（Ana 2）连接到一个单一的调控途径中。任何这些蛋白质的过表达诱导组装的多个子中心粒。细胞周期控制程序如何调节这些关键蛋白质以限制子中心粒组装仅为单一事件尚不清楚。在这里，我们假设，细胞周期调控，包括后期促进复合物（APC）泛素连接酶，控制破坏的Plk 4/Asl/Ana 2复合物，改变其模式/activit母亲中心粒，并产生一个单一的网站的女儿组装。我们的初步数据表明：1）中心粒上的Plk 4模式在有丝分裂过程中发生变化：首先覆盖整个母中心粒，然后通过有丝分裂退出被修剪成单个不对称点; 2）Plk 4和Ana 2是APC靶点; 3）磷酸化保护这些蛋白质的亚群免于降解; 4）Asl是稳定有丝分裂Plk 4的Plk 4底物。在本提案中，我们 将（1）确定中心粒上Plk 4定位的APC调节是否限定了子代中心粒组装的位点，（2）确定Plk 4和Asl之间的功能性相互作用，其有助于限定中心粒组装的单个位点，和（3）确定Ana 2是否调节Plk 4活化和Ana 2是否由APC控制。这些研究将提供一个分子过程，管理中心粒复制的保真度机制的见解。在分子水平上理解这一过程将指导未来的研究，以探索肿瘤发生和其他中心粒相关疾病中中心粒扩增的病因。