STRUCTURE AND FUNCTION OF YEAST H1 HISTONES

一般认为酵母属 酿酒酵母不含H1组蛋白， 参与核小体纤维的凝聚 高等真核生物 我们发现，通过执行功能性 HI组蛋白的测定，酵母染色质由 核小体含有H1组蛋白和168 碱基对（bp）DNA。 此外，H1样活性可以是 用0.6M NaCl提取，并且可以组装染色体 从H1耗尽的染色质在0.6 M的存在下 提取物 在本申请中，我们建议调查 酵母中H1组蛋白的结构和功能 使用遗传学和生物化学方法。 我们将净化 来自0.6 M NaCl提取物和克隆的酵母H1组蛋白 并测序H1基因。 推导的氨基酸序列 将与哺乳动物的H1组蛋白进行比较， H1结构的相似性和差异将是 测定 酵母染色体中的H1基因将 通过用标记基因替换H1基因而失活， H1组蛋白在酵母中的生物学功能将是 通过对H1耗竭影响的研究进行了调查 组蛋白对细胞周期、基因转录和染色质的影响 结构 H1在酵母缩合中的作用 核小体纤维将通过观察离子 核小体纤维强度依赖性折叠 电子显微镜，以及H1的重要区域， 核小体纤维凝聚将通过 H1基因的部分缺失。 Hi的磷酸化 在细胞周期中，将进行研究。 将 确定Hi是否确实是细胞周期的底物 控制蛋白激酶（CDC 28）在体内，因为其他人有 建议道。

It has been generally accepted that yeast Saccharomyces cerevisiae do not contain H1 histones, the histones which are involved in the condensation of the nucleosome fibre in higher eucaryotes. We discovered, by carrying out functional assays for HI histones, that yeast chromatin consists of chromatosomes, the nucleosomes containing H1 histones and 168 base pair (bp) DNA. Furthermore, the H1-like activity can be extracted by 0.6 M NaC1, and chromatosomes can be assembled from H1-depleted chromatin in the presence of the 0.6 M extract. In this application, we propose to investigate the structure and function of the H1 histones in yeast with the use of genetics and biochemical approaches. We will purify the yeast H1 histones from the 0.6 M NaC1 extract and clone and sequence the H1 gene(s). The deduced amino acid sequence will be compared with those of mammalian H1 histones, and the similarity and differences in the structure of H1 will be determined. The H1 gene in the yeast chromosome will be inactivated by replacement of the H1 gene with marker genes, and the biological function of H1 histones in yeast will be investigated by studies on the effects of depletion of H1 histones on the cell cycle, gene transcription, and chromatin structure. The role of H1 in the condensation of yeast nucleosome fibre will be investigated by observation of ionic strength-dependent folding of nucleosome fibre under an electron microscope, and the regions of the H1 important in the condensation of nucleosome fibre will be investigated by partial deletions of the H1 gene. The phosphorylation of Hi during the cell cycle will be investigated. It will be determined if Hi is indeed the substrate for the cell cycle control protein kinase (CDC28) in vivo as others have suggested.