A functional role for nucleosomes in the repression of a yeast promoter.
A functional role for nucleosomes in the repression of a yeast promoter.
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DOI：
10.1002/j.1460-2075.1991.tb07957.x
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发表时间：
1991-02
期刊：
影响因子：
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通讯作者：
Christian Straka;W. Horz
中科院分区：
文献类型：
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作者：
Christian Straka;W. Horz
PHO 5基因在S.先前显示，酿酒酵母的启动子中的四个定位的核小体的去除伴随着从启动子中去除。为了评估核小体在基因激活级联中的作用，切除对应于这些核小体之一的DNA。在其位置插入两个相同长度的外源DNA片段：来自非洲绿色猴α-卫星DNA的片段，已知其以高度特异性的方式与组蛋白结合，以产生独特定位的核小体，或者，来自pBR 322 DNA的片段。将启动子构建体与着丝粒质粒上的lacZ基因融合并转化到酵母细胞中。卫星片段形成核小体，在诱导条件下持续存在。同时，PHO 5启动子的诱导作用几乎被消除。当检测含有35 - 100 bp卫星片段的各种亚片段时，发现它们都降低了启动子的诱导性，然而，完全阻遏需要全长分子。相比之下，pBR片段使启动子弱组成型，并且诱导进行到甚至比缺乏插入物的启动子更高的水平。染色质结构的分析揭示了在非诱导条件下pBR片段上的核小体，其在诱导后被去除。可以得出结论，启动子处组蛋白-DNA相互作用的质量对基因的调控做出了内在贡献。
Induction of the PHO5 gene in S. cerevisiae was previously shown to be accompanied by the removal of four positioned nucleosomes from the promoter. In order to assess the role of nucleosomes in the cascade of gene activation, DNA corresponding to one of these nucleosomes was excised. In its place two foreign DNA segments of the same length were inserted: a fragment from the African green monkey alpha‐satellite DNA which is known to associate with histones in a highly specific fashion to give a uniquely positioned nucleosome or, alternatively, a fragment derived from pBR322 DNA. The promoter constructs were fused to the lacZ gene on centromere plasmids and transformed into yeast cells. The satellite fragment formed a nucleosome which persisted under inducing conditions. At the same time the inducibility of the PHO5 promoter was virtually abolished. When various subfragments containing between 35 and 100 bp of the satellite segment were tested, they were all found to decrease the inducibility of the promoter, full repression required the full length molecule, however. In contrast, the pBR fragment made the promoter weakly constitutive, and induction proceeded to levels even higher than with a promoter lacking an insert. Analysis of the chromatin structure reveals a nucleosome on the pBR segment at noninducing conditions which is removed upon induction. It is concluded that the quality of the histone‐DNA interactions at the promoter makes an intrinsic contribution to the regulation of the gene.