STRUCTURE AND FUNCTION OF YEAST H1 HISTONES

人们普遍认为，酵母菌 酿酒酵母不包含H1组蛋白，这些组蛋白是 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 对HI组蛋白的检测，酵母染色质包括 染色体，含有H1组蛋白和168的核小体 碱基对(BP)DNA。此外，类H1活动可以是 用0.6M氯化钠提取，可组装染色体 在0.6M存在的情况下从H1耗尽的染色质 提取。在此应用程序中，我们建议调查 酵母组蛋白H1的结构和功能 遗传学和生物化学方法的使用。我们将净化 0.6M氯化钠提取和克隆酵母H1组蛋白 并对H1基因进行测序(S)。推导出的氨基酸序列 将与哺乳动物的H1组蛋白进行比较，并且 H1结构的相似和不同之处将是 下定决心。酵母染色体中的H1基因将是 通过用标记基因替换H1基因来使其失活， 而酵母中H1组蛋白的生物学功能将是 通过对H1耗竭的影响的研究进行调查 组蛋白对细胞周期、基因转录和染色质的影响 结构。H1在酵母缩合反应中的作用 核小体纤维将通过离子观察进行研究 核小体纤维在压力作用下的强度依赖性折叠 电子显微镜，以及H1的重要区域 核小体纤维的凝聚将通过 H1基因的部分缺失。Hi的磷酸化 在细胞周期中将被调查。会是 确定Hi是否确实是细胞周期的底物 体内的控制蛋白激酶(CDC28) 建议。

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.