Han M, Kim U J, Kayne P, Grunstein M
Molecular Biology Institute, University of California, Los Angeles 90024.
EMBO J. 1988 Jul;7(7):2221-8. doi: 10.1002/j.1460-2075.1988.tb03061.x.
We have previously constructed a yeast strain (UKY403) whose sole histone H4 gene is under control of the GAL1 promoter. This yeast arrests in G2 upon glucose treatment as a result of histone H4 depletion. The yeast PHO5 gene contains phase nucleosomes covering promoter (UAS) sequences in the PHO5 repressed state and it has been suggested that nucleosomes prevent the binding of positively acting factors to these UAS sequences. Using UKY403 we examined the length of polynucleosomes and nucleosome phasing in the PHO5 upstream region by the use of micrococcal nuclease and indirect end-labeling. It was found that glucose arrest led to a severe disruption in PHO5 chromatin structure and that most nucleosomes had their position altered or were lost from the PHO5 promoter region. Cell undergoing nucleosome depletion synthesized large quantities of accurate PHO5 transcripts even under repressive, high inorganic phosphate conditions. Histone H4 depletion did not appear to affect the repression or activation of another inducible yeast gene, CUP1. Arrest with landmarks in early G1 (in the cell division cycle mutant cdc28) or in various stages of G2 (in cdc15, cdc17 and cdc20) does not activate PHO5; nor does arrest due to chromosome topology changes (in top2 or the top1top2 topoisomerase mutants). cdc14, which has its arrest landmark at a similar point in the cell cycle as cdc15, does derepress PHO5. However, since it also leads to derepression of CUP1 it is probably functioning through an independent mechanism. Therefore, our data suggest that nucleosomes regulate PHO5 transcription.
我们之前构建了一种酵母菌株（UKY403），其唯一的组蛋白H4基因受GAL1启动子控制。由于组蛋白H4缺失，这种酵母在葡萄糖处理后会停滞在G2期。酵母PHO5基因在PHO5处于抑制状态时含有覆盖启动子（UAS）序列的阶段性核小体，有人认为核小体会阻止正向作用因子与这些UAS序列结合。利用UKY403，我们通过微球菌核酸酶和间接末端标记法研究了PHO5上游区域多核小体的长度和核小体相位。结果发现，葡萄糖诱导的停滞导致PHO5染色质结构严重破坏，大多数核小体的位置发生改变或从PHO5启动子区域消失。即使在抑制性的高无机磷酸盐条件下，经历核小体缺失的细胞仍能合成大量准确的PHO5转录本。组蛋白H4缺失似乎并不影响另一个可诱导酵母基因CUP1的抑制或激活。在G1早期（细胞分裂周期突变体cdc28）或G2的各个阶段（cdc15、cdc17和cdc20）停滞时不会激活PHO5；由于染色体拓扑结构变化（top2或top1top2拓扑异构酶突变体）导致的停滞也不会激活PHO5。cdc14在细胞周期中的停滞标志点与cdc15相似，它确实能解除对PHO5的抑制。然而，由于它也会导致CUP1的去抑制，所以它可能是通过一种独立的机制发挥作用。因此，我们的数据表明核小体调节PHO5的转录。
