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首页> 外文期刊>Molecular and Cellular Biology >Constitutive repression and nuclear factor I-dependent hormone activation of the mouse mammary tumor virus promoter in Saccharomyces cerevisiae.
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Constitutive repression and nuclear factor I-dependent hormone activation of the mouse mammary tumor virus promoter in Saccharomyces cerevisiae.

机译：酿酒酵母中小鼠乳腺肿瘤病毒启动子的组成型抑制和核因子I依赖性激素激活。

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To study the influence of various transactivators and the role of nucleosomal structure in gene regulation by steroid hormones, we have introduced mouse mammary tumor virus (MMTV) promoter sequences along with expression vectors for the glucocorticoid receptor (GR) and nuclear factor I (NFI) in Saccharomyces cerevisiae, an organism amenable to genetic manipulation. Both in the context of an episomal multicopy vector and in a centromeric single-copy plasmid, the MMTV promoter was virtually silent in the absence of inducer, even in yeast strains expressing GR and NFI. Induction was optimal with deacylcortivazol and required both GR and NFI. The transactivation function AF1 in the N-terminal half of GR is required for ligand-dependent induction and acts constitutively in truncated GR lacking the ligand binding domain. A piece of the MMTV long terminal repeat extending from -236 to +111 is sufficient to position a nucleosome, B, over the regulatory region of the promoter from -45 to -190 and another nucleosome over the transcription start region. The rotational orientation of the DNA on the surface of nucleosome B is the same as that previously found in animal cells and in reconstitution experiments. This orientation is compatible with binding of GR to two sites, while it should preclude binding of NFI and hence be responsible for constitutive repression. Upon ligand induction, there is no major chromatin rearrangement, but the proximal linker DNA, including the TATA box, becomes hypersensitive to nucleases. The transcriptional behavior of the MMTV promoter was unaffected by deletions of the genes for zuotin or SIN1/SPT2, two proteins which have been claimed to assume some of the functions of linker histones. Thus, despite the lack of histone H1, yeast cells could be a suitable system to study the contribution of nucleosomal organization to the regulated expression of the MMTV promoter.
机译：为了研究各种反式激活因子的影响以及核糖体结构在类固醇激素基因调控中的作用，我们引入了小鼠乳腺肿瘤病毒（MMTV）启动子序列以及糖皮质激素受体（GR）和核因子I（NFI）的表达载体在酿酒酵母中，它是一种易于进行基因操作的生物。在游离型多拷贝载体的背景下和在着丝粒单拷贝质粒中，MMTV启动子在没有诱导剂的情况下实际上是沉默的，即使在表达GR和NFI的酵母菌株中也是如此。用去酰基Cortivazol诱导是最佳的，并且需要GR和NFI。 GR的N末端一半的反式激活功能AF1是依赖配体的诱导所必需的，并且在缺乏配体结合域的截短GR中起组成性作用。一段从-236延伸到+111的MMTV长末端重复序列足以将一个核小体B定位在启动子从-45到-190的调控区域上，而另一个核小体则位于转录起始区域之上。 DNA在核小体B表面的旋转方向与先前在动物细胞和重组实验中发现的方向相同。这种方向与GR与两个位点的结合是兼容的，同时它应排除NFI的结合，因此对组成型抑制负责。配体诱导后，没有主要的染色质重排，但包括TATA盒在内的近端接头DNA对核酸酶变得高度敏感。 MMTV启动子的转录行为不受zuotin或SIN1 / SPT2基因缺失的影响，这两个蛋白据称具有接头组蛋白的某些功能。因此，尽管缺乏组蛋白H1，酵母细胞仍可能是研究核小体组织对MMTV启动子调控表达的贡献的合适系统。

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