Vicent Guillermo P, Meliá María J, Beato Miguel
Institüt für Molekularbiologie und Tumorforschung (IMT), Philipps-Universität, Emil-Mannkoppf-Str. 2, D-35033, Marburg, Germany.
J Mol Biol. 2002 Nov 29;324(3):501-17. doi: 10.1016/s0022-2836(02)01101-4.
Packaging of mouse mammary tumor virus (MMTV) promoter sequences in nucleosomes modulates access of DNA binding proteins and influences the interaction among DNA bound transcription factors. Here we analyze the binding of histone H1 to MMTV mononucleosomes assembled with recombinant histones and study its influence on nucleosome structure and stability as well as on progesterone receptor (PR) binding to the hormone responsive elements (HREs). The MMTV nucleosomes can be separated into three main populations, two of which exhibited precise translational positioning. Histone H1 bound preferentially to the 5' distal nucleosomal DNA protecting additional 27-28 nt from digestion by micrococcal nuclease. Binding of histone H1 was unaffected by prior crosslinking of protein and DNA in nucleosomes with formaldehyde. Neither the translational nor the rotational nucleosome positioning was altered by histone H1 binding, but the nucleosomes were stabilized as judged by the kinetics of nuclease cleavage. Unexpectedly, binding of recombinant PR to the exposed distal HRE-I in nucleosomes was enhanced in the presence of histone H1, as demonstrated by band shift and footprinting experiments. This enhanced PR affinity may contribute to the reported positive effect of histone H1 on the hormonal activation of MMTV reporter genes.
小鼠乳腺肿瘤病毒（MMTV）启动子序列在核小体中的包装调节了DNA结合蛋白的可及性，并影响了DNA结合转录因子之间的相互作用。在此，我们分析了组蛋白H1与用重组组蛋白组装的MMTV单核小体的结合，并研究了其对核小体结构和稳定性以及对孕激素受体（PR）与激素反应元件（HREs）结合的影响。MMTV核小体可分为三个主要群体，其中两个表现出精确的平移定位。组蛋白H1优先结合到5'远端核小体DNA上，保护另外27 - 28个核苷酸不被微球菌核酸酶消化。组蛋白H1的结合不受核小体中蛋白质和DNA先前用甲醛交联的影响。组蛋白H1的结合既不改变平移也不改变旋转核小体定位，但从核酸酶切割动力学判断，核小体得到了稳定。出乎意料的是，如凝胶迁移和足迹实验所示，在组蛋白H1存在的情况下，重组PR与核小体中暴露的远端HRE-I的结合增强。这种增强的PR亲和力可能有助于报道的组蛋白H1对MMTV报告基因激素激活的积极作用。
