Crotti Luciana B, Basrai Munira A
Genetics Branch, National Cancer Institute, Bethesda, MD 20889-5105, USA.
EMBO J. 2004 Apr 21;23(8):1804-14. doi: 10.1038/sj.emboj.7600161. Epub 2004 Apr 1.
The kinetochore (centromeric DNA and associated proteins) mediates the attachment of chromosomes to the mitotic spindle apparatus and is required for faithful chromosome transmission. We established that evolutionarily conserved Saccharomyces cerevisiae SPT4, previously identified in genetic screens for defects in chromosome transmission fidelity (ctf), encodes a new structural component of specialized chromatin at kinetochores and heterochromatic loci, with roles in kinetochore function and gene silencing. Using chromatin immunoprecipitation assays (ChIP), we determined that kinetochore proteins Ndc10p, Cac1p, and Hir1p are required for the association of Spt4p to centromeric (CEN) loci. Absence of functional Spt4p leads to altered chromatin structure at the CEN DNA and mislocalization of the mammalian CENP-A homolog Cse4p to noncentromeric loci. Spt4p associates with telomeres (TEL) and HMRa loci in a Sir3p-dependent manner and is required for transcriptional gene silencing. We show that a human homolog of SPT4 (HsSPT4) complements Scspt4-silencing defects and associates with ScCEN DNA in an Ndc10p-dependent manner. Our results highlight the evolutionary conservation of pathways required for genome stability in yeast and humans.
动粒（着丝粒DNA及相关蛋白）介导染色体与有丝分裂纺锤体装置的附着，是染色体准确传递所必需的。我们确定，在染色体传递保真度（ctf）缺陷的遗传筛选中先前鉴定出的进化保守的酿酒酵母SPT4，编码动粒和异染色质位点处特殊染色质的一种新结构成分，在动粒功能和基因沉默中发挥作用。通过染色质免疫沉淀测定法（ChIP），我们确定动粒蛋白Ndc10p、Cac1p和Hir1p是Spt4p与着丝粒（CEN）位点结合所必需的。功能性Spt4p的缺失会导致CEN DNA处染色质结构改变，以及哺乳动物CENP - A同源物Cse4p错定到非着丝粒位点。Spt4p以Sir3p依赖的方式与端粒（TEL）和HMRa位点结合，是转录基因沉默所必需的。我们表明，SPT4的人类同源物（HsSPT4）可弥补Scspt4的沉默缺陷，并以Ndc10p依赖的方式与ScCEN DNA结合。我们的结果突出了酵母和人类中基因组稳定性所需途径的进化保守性。
