The structural analysis of centromere specific nucleosome

在真核生物中，基因组DNA在S期复制，并在M期平均分裂成两个子细胞。着丝粒是染色体上的一个独特区域，在这一分裂过程中起着重要的作用。着丝粒的形成是染色体分离所必需的。但着丝粒区域是如何形成和区别于其他区域的，目前尚不清楚。鉴定了一些特异定位于着丝粒区域的蛋白质。CENP-A是组蛋白H3的变异体，也定位于着丝粒区域。CENP-A被组装到着丝粒的染色质中，并被认为产生独特的染色质特征。在本研究中，我们对CENP-A核小体进行了结构分析，并用X射线结晶学方法测定了CENP-A核小体的晶体结构。结构测定表明，CENP-A核小体含有两个组蛋白H_2A、H_2B、H_4和CENP-A，DNA以左手方向缠绕在组蛋白八聚体周围。令人惊讶的是，在CENP-A-核小体中，只有121个碱基的DNA可见，而DNA两端的13个碱基是灵活的。CENP-A和H3-核小体的结构比较表明，CENP-A的loop1区不同于典型的H3，这是因为插入了两个额外的氨基酸残基。体内分析表明，这些氨基酸对于将CENP-A保留在着丝粒区域是重要的。

In eukaryotes, genomic DNA is duplicated during S phase, and is equally divided into two daughter cells during M phase. Centromere, which is one of the distinct regions in chromosome, plays important rules in this division. Centromere formation is necessary for chromosome segregation. But, it is not known how centromere region is formed and distinguished from other region. Some proteins, which localize specifically at centromere region, are identified. CENP-A, a histone H3 variant, also localize specifically at centromere region. CENP-A is assembled into the chromatin of centromeres and is thought to generate unique chromatin features. In this study, we performed structural analysis of the CENP-A nucleosome and determined crystal structure of CENP-A nucleosome by X-ray crystallography. Determined structure revealed that CENP-A nucleosome contains two each of histones H2A, H2B, H4 and CENP-A and DNA is wrapped around histone octamer in a left-hand orientation. Surprisingly, only 121 bp of DNA are visible and 13 bp from both ends of DNA are flexible in the CENP-A-nucleosome. Structural comparison of the CENP-A and H3-nucleosomes revealed that the loop1 region of CENP-A is different from canonical H3 because of insertion of two extra amino acid residues. In vivo analysis revealed these amino acids are important to retain CENP-A at centromere region.