STRUCTURAL ANALYSIS OF AN EUKARYOTIC CENTROMERE

真核生物染色体的着丝粒是染色体沿着 染色质纤维在染色体运动中起着重要作用， 细胞分裂 从酵母中分离的着丝粒DNA序列， 酿酒酵母，使外源DNA引入酵母， 在细胞分裂过程中起着普通酵母染色体的作用。 我们将 研究着丝粒DNA序列如何与染色质成分相互作用 在细胞核中，包括核蛋白和微管， 形成一个有功能的着丝粒单位。 我们会在地图上 核酸酶切割位点，并将测量DNA双链体的螺旋 在着丝粒染色质内决定着丝粒的组织 DNA、组蛋白和非组蛋白。 这些蛋白质与 将通过直接分离着丝粒DNA来鉴定 胞内着丝粒质粒蛋白复合物。 结合的蛋白质 将通过过滤器根据其DNA结合特性来表征 结合试验及其分子异质性（凝胶法） 电泳 然后，我们将研究这些蛋白质如何与 着丝粒DNA和微管产生功能性着丝粒 单位 随着着丝粒特异性DNA结合蛋白的分离， 最终应该能够重建着丝粒纺锤纤维 复合物体外培养。 了解控制组织的因素 着丝粒的可能提供基本原则， 细胞分裂的分子机制。 细胞分裂是一个核心过程 生命有机体。 细胞分裂控制的变化可能是一种 导致产生异常细胞的重要机制 包括癌细胞。 着丝粒异常，过早 着丝粒分离和染色体不稳定的情况下， Roberts' - SC短肢畸形、共济失调毛细血管扩张症和一些其他 综合征 对控制细胞分裂的因素的理解可能 是进一步了解这些功能障碍的基础。

Centromeres of eukaryotic chromosomes are specific regions along the chromatin fiber that play a fundamental role in chromosome movement during cell division. Centromere DNA sequences isolated from the yeast, Saccharomyces cerevisiae, enable foreign DNA introduced into yeast to function as ordinary yeast chromosomes during cell division. We will examine how the centromere DNA sequence interacts with chromatin components in the cell nucleus, including nuclear proteins and microtubules, to give rise to a functional centromeric unit. We will map the location of nuclease cleavage sites and will measure the coiling of the DNA duplex within centromeric chromatin to determine the organization of centromere DNA, histones and nonhistone proteins. The proteins that are associated with centromere DNA in vivo will be identified by direct isolation of intracellular centromere plasmid-protein complexes. The bound proteins will be characterized in terms of their DNA-binding properties by a filter binding assay and for their molecular heterogeneity by gel electrophoresis. We will then examine how these proteins interact with centromere DNA and microtubules to give rise to a functional centromeric unit. With the isolation of centromere specific DNA-binding proteins we should eventually be able to reconstitute the centromere spindle-fiber complex in vitro. Knowledge of the factors that control the organization of the centromere may provide fundamental principles governing the molecular mechanisms of cell division. Cell division is a central process of living organisms. Changes in the control of cell division may be an important mechanism that results in the production of aberrant cell populations, including cancer cells. Centromeric abnormalities, premature centromere separation and chromosome instability have been found in cases of Roberts' - SC phocomelia, ataxia telangiectasia, and a number of other syndromes. An understanding of the factors controlling cell division may be fundamental for the further understanding of these dysfunctions.