Keith K C, Baker R E, Chen Y, Harris K, Stoler S, Fitzgerald-Hayes M
Department of Biochemistry and Molecular Biology, Program in Molecular and Cellular Biology, University of Massachusetts at Amherst, Amherst, Massachusetts 01003, USA.
Mol Cell Biol. 1999 Sep;19(9):6130-9. doi: 10.1128/MCB.19.9.6130.
Cse4p is a variant of histone H3 that has an essential role in chromosome segregation and centromere chromatin structure in budding yeast. Cse4p has a unique 135-amino-acid N terminus and a C-terminal histone-fold domain that is more than 60% identical to histone H3 and the mammalian centromere protein CENP-A. Cse4p and CENP-A have biochemical properties similar to H3 and probably replace H3 in centromere-specific nucleosomes in yeasts and mammals, respectively. In order to identify regions of Cse4p that distinguish it from H3 and confer centromere function, a systematic site-directed mutational analysis was performed. Nested deletions of the Cse4p N terminus showed that this region of the protein contains at least one essential domain. The C-terminal histone-fold domain of Cse4p was analyzed by changing Cse4p amino acids that differ between Cse4p and H3 to the analogous H3 residues. Extensive substitution of contiguous Cse4p residues with H3 counterparts resulted in cell lethality. However, all large lethal substitution alleles could be subdivided into smaller viable alleles, many of which caused elevated rates of mitotic chromosome loss. The results indicate that residues critical for wild-type Cse4p function and high-fidelity chromosome transmission are distributed across the entire histone-fold domain. Our findings are discussed in the context of the known structure of H3 within the nucleosome and compared with previous results reported for CENP-A.
Cse4p是组蛋白H3的一种变体，在芽殖酵母的染色体分离和着丝粒染色质结构中起关键作用。Cse4p有一个独特的135个氨基酸的N端和一个C端组蛋白折叠结构域，该结构域与组蛋白H3以及哺乳动物着丝粒蛋白CENP - A的同源性超过60%。Cse4p和CENP - A具有与H3相似的生化特性，可能分别在酵母和哺乳动物的着丝粒特异性核小体中取代H3。为了确定Cse4p中使其区别于H3并赋予着丝粒功能的区域，进行了系统的定点突变分析。Cse4p N端的嵌套缺失表明该蛋白区域至少包含一个必需结构域。通过将Cse4p和H3之间不同的氨基酸替换为类似的H3残基，对Cse4p的C端组蛋白折叠结构域进行了分析。用H3对应残基广泛取代相邻的Cse4p残基导致细胞致死。然而，所有大的致死性取代等位基因都可细分为较小的存活等位基因，其中许多导致有丝分裂染色体丢失率升高。结果表明，对野生型Cse4p功能和高保真染色体传递至关重要的残基分布在整个组蛋白折叠结构域。我们的研究结果在核小体中H3已知结构的背景下进行了讨论，并与先前报道的CENP - A的结果进行了比较。
