Centromere identity and function

描述（由申请人提供）：该提案关注遗传学中的一个基本问题：染色体及其携带的基因如何在细胞分裂时适当地分离到子细胞中？这一过程中的缺陷导致基因组的不均匀分配，这反过来又导致非整倍性。如果这种情况发生在配子发生期间，这种类型的遗传灾难会导致自然流产或后代出生缺陷。染色体缺失：由于有丝分裂细胞的错误沿着肿瘤抑制基因的突变而导致的单个染色体的增加或丢失 基因或癌基因有助于肿瘤形成和发展的早期事件，并且非整倍性是几乎所有晚期肿瘤的标志。控制其精确分离的染色体元件是着丝粒。分子遗传学和人类患者数据已经定义了着丝粒身份，而不是通过特定的DNA序列，因为着丝粒？在大多数正常的着丝粒上发现的卫星重复既不必要也不足以确定功能性着丝粒的位置。相反，流行的观点是，着丝粒的身份是表观遗传学定义的，我们的长期目标是了解这种表观遗传着丝粒标记的基础和用于建立和维护的机制。代表表观遗传标记的主要候选者是CENP-A，一种在功能性着丝粒处组装成核小体的组蛋白H3变体。本提案中的具体研究将解决以下问题：CENP-A如何在着丝粒处产生表观遗传标记？CENP-A掺入多核小体阵列对其高阶折叠有什么影响？含有CENP-A的核小体是否直接募集一组特定的着丝粒因子？CENP-A对着丝粒有选择性亲和力吗？卫星DNA？CENP-A装载到着丝粒DNA上的机制是什么？什么样的细胞蛋白质通过其着丝粒靶向结构域识别CENP-A？ 使用生物化学重建，生物物理学，分子遗传学和细胞生物学方法的组合，我们提出了以下具体目标： 1.确定CENP-A掺入着丝粒核小体的结果 2.定义CENP-A染色质在着丝粒组装的要求 总之，这些研究有望提供有价值的见解，表观遗传机制，指定和维持染色体上的着丝粒的位置。

DESCRIPTION (provided by applicant): This proposal is focused on a fundamental question in genetics: How are chromosomes and the genes they carry appropriately segregated to the daughter cells at cell division? Defects in this process result in uneven partitioning of the genome that, in turn, causes aneuploidy. If this happens during gametogenesis, this type of genetic catastrophe results in spontaneous abortion or birth defects in resulting offspring. The gain or loss of an individual chromosome resulting from errors in mitotic cell division along with mutation of tumor suppressor genes or oncogenes contributes to early events in tumor formation and progression, and aneuploidy is a hallmark of almost all late stage tumors. The chromosomal element that controls its accurate segregation is the centromere. Molecular genetic and human patient data has defined centromere identity not by a particular DNA sequence, since centromeric ?-satellite repeats found at most normal centromeres are neither necessary nor sufficient to specify the location of a functioning centromere. Rather, the prevailing view is that centromere identity is defined epigenetically, and our long-term goal is to understand the basis for this epigenetic centromere mark and the mechanisms used for its establishment and maintenance. The leading candidate to represent the epigenetic mark is CENP-A, a histone H3 variant that assembles into nucleosomes at functional centromeres. Specific research in this proposal will address the following questions: How does CENP-A generate an epigenetic mark at the centromere? What impact does the incorporation of CENP-A into polynucleosome arrays have on their higher-order folding? Does the CENP-A-containing nucleosome directly recruit a specific set of centromere factors? Is there selective affinity of CENP-A for centromeric ?-satellite DNA? What is the mechanism for loading CENP-A onto centromeric DNA? What cellular proteins recognize CENP-A via its centromere targeting domain? Using a combination of biochemical reconstitution, biophysics, molecular genetics, and cell biology approaches we propose the following specific aims: 1. Determine the consequences of incorporation of CENP-A into centromeric nucleosomes 2. Define the requirements for CENP-A chromatin assembly at centromeres Together, these studies promise to provide valuable insight into the epigenetic mechanisms that specify and maintain the location of the centromere on the chromosome.