Mechanistic Basis for CENP-32 Mediated Regulation of Cell Division

在人体内，每天有数万亿个细胞进行分裂。在每次分裂过程中，以染色体形式存在的遗传信息需要平等和相同地分配给新形成的子细胞。染色体的分布是由一种称为有丝分裂纺锤体的复杂机制实现的。在人类中，有丝分裂纺锤体是由称为微管的丝状网络形成的，微管由称为中心体的细胞器组织。在细胞分裂过程中，细胞具有两个中心体，它们彼此远离，并协调微管网络以建立有丝分裂纺锤体。中心体形成有丝分裂纺锤体的两端，微管网络在此汇聚，称为纺锤体极。中心体与纺锤体极的物理连接对于将染色体和中心体准确地分配到子细胞中至关重要。染色体和/或中心体的缺陷分布与几种人类健康疾病如癌症、小头畸形和原始侏儒症有关。因此，了解中心体是如何附着在纺锤体两极的，对于更好地了解相关的医学状况和找到可能的治疗方法是很重要的。拟议的工作集中在一种名为CENP-32的人类必需蛋白质上，当从细胞中取出时，会导致中心体从纺锤体极点脱离，影响有丝分裂纺锤体的完整性。CENP-32突变也在患有神经发育障碍的患者中发现，如小头畸形，癫痫发作和发育迟缓。在两极缺乏中心体的有丝分裂纺锤体不能准确地分配染色体，因此将导致具有不适当的染色体和/或中心体数目的子细胞，这通常与癌症和发育障碍有关。蛋白质通过与包括蛋白质和核酸（RNA和DNA）的生物分子相互作用和/或修饰生物分子来发挥其功能。蛋白质通过其三维结构获得其功能，这为它们提供了与其他生物分子相互作用/修饰其他生物分子的能力。为了了解CENP-32，一种可能的RNA结合蛋白，如何确保中心体与纺锤体极的物理连接，我们提出：（1）研究CENP-32的结构并鉴定其进行的RNA修饰，（2）鉴定与CENP-32相互作用的蛋白质/RNA，（3）描述CENP-32的RNA修饰活性如何促进中心体-纺锤体极附着，以及（4）评估CENP-32患者突变如何影响CENP-32的活性和功能。这项研究的结果将促进我们对中心体如何帮助构建完整的有丝分裂纺锤体的理解，该纺锤体对于产生具有正确遗传信息的子细胞至关重要。CENP-32是细胞存活所必需的蛋白质，因此，拟议研究的结果也将为探索阻断癌细胞中CENP-32功能和纠正相关神经发育障碍患者中CENP-32缺陷的可能性铺平道路。

In the human body, trillions of cells undergo division every day. During each division, the genetic information which is in the form of chromosomes need to be equally and identically distributed to the newly formed daughter cells. Distribution of chromosomes is achieved by an elaborate machinery called the mitotic spindle. In humans, the mitotic spindle is formed by a filamentous network called microtubules which are organised by an organelle called centrosome. During cell division cells possess two centrosomes that move away from each other and orchestrate microtubule network to establish the mitotic spindle. The centrosomes form the opposite ends of the mitotic spindle where microtubule networks converge and are called spindle poles. Physical attachment of centrosomes to the spindle poles is crucial for distributing chromosomes and centrosomes accurately to the daughter cells. Defective distribution of chromosomes and/or centrosomes are associated with several human health disorders such as cancer, microcephaly and primordial dwarfism. Hence understanding how centrosomes are attached to the spindle poles is important to better understand the related medical conditions and to find a possible cure. The proposed work focuses on an essential human protein called CENP-32, which when removed from cells results in the detachment of centrosomes from spindle poles affecting the integrity of the mitotic spindle. CENP-32 mutations have also been found in patients with neurodevelopmental disorders such as microcephaly, seizures and developmental delays. The mitotic spindle lacking centrosomes at the poles is not capable of distributing the chromosomes accurately and as a consequence will result in daughter cells with inappropriate chromosome and/or centrosome numbers, a condition often associated with cancer and developmental disorders. Proteins exert their function by interacting with and/or modifying biomolecules including proteins and nucleic acids (RNA and DNA). Proteins acquire their function through their three-dimensional structure which provides them their ability to interact with/modify other biomolecules. To understand how CENP-32, a likely RNA binding protein, ensures the physical attachment of centrosomes to spindle poles, we propose: (1) to study the structure of CENP-32 and identify the RNA modifications it makes, (2) to identify the proteins/RNA that CENP-32 interacts with, (3) to delineate how RNA modifying activity of CENP-32 facilitate centrosome-spindle pole attachment, and (4) assess how CENP-32 patient mutations affect CENP-32 activity and function.The outcome of this research will advance our understanding of how centrosomes help build an intact mitotic spindle essential for generating daughter cells with correct genetic information. CENP-32 is a protein essential for cell survival, hence the outcome of the proposed research will also pave way for exploring the possibility of blocking CENP-32 function in cancer cells and rectifying CENP-32 defect in patients with associated neurodevelopmental disorders.