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首页> 外文期刊>Molecular and Cellular Biology >Detection of two chromatin proteins which bind specifically to the 5'-flanking region of the rat prolactin gene.
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Detection of two chromatin proteins which bind specifically to the 5'-flanking region of the rat prolactin gene.

机译：检测到两个与大鼠催乳素基因的5'侧翼区域特异性结合的染色质蛋白。

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We employed a protein gel blotting procedure to search for nuclear proteins from rat pituitary cells that bind preferentially to the 5'-flanking region of the rat prolactin gene. By gel blots of chromatin proteins from GH3 rat pituitary tumor cells with a 32P-labeled prolactin genomic clone, we detected two major binding proteins with molecular weights of approximately 44,000 and 48,000, designated NP44 and NP48, respectively. Both NP44 and NP48 are minor chromatin proteins which are extracted at low salt concentrations (0.4 M NaCl) and exhibit a range of slightly acidic isoelectric variants. NP44 and NP48 were detected at similar levels in chromatin extracts of GH3 cells, the prolactin-negative GC cell variant of the GH3 cells, and normal rat pituitary tissue. Considerably lower levels of these two proteins were found in chromatin extracts from rat liver and rat C6 glial cells. NP44 and NP48 exhibit DNA sequence specificity, as evidenced by their strong binding to the upstream flanking region of the prolactin gene, but only very weak binding to plasmid DNA, rat prolactin or growth hormone cDNAs, or upstream flanking regions of two other rat genes. By analyzing subclones of a rat prolactin genomic clone, we established that NP44 and NP48 bind to at least two sites, which are located between 0.4 and 2.0 kilobases (region I) and between 2.0 and 4.8 kilobases (region II) upstream of the transcription initiation site. These findings are discussed in the context of a possible functional association between the strong binding of NP44 and NP48 to the prolactin 5'-flanking region and pituitary-specific expression of the prolactin gene.
机译：我们采用蛋白质凝胶印迹程序从大鼠垂体细胞中搜索优先结合大鼠催乳素基因5'侧翼区的核蛋白。通过带有32P标记催乳素基因组克隆的GH3大鼠垂体肿瘤细胞的染色质蛋白的凝胶印迹，我们检测到了两种主要的结合蛋白，分子量分别约为44,000和48,000，分别命名为NP44和NP48。 NP44和NP48都是次要的染色质蛋白，它们以低盐浓度（0.4 M NaCl）提取，并显示出一系列的弱酸性等电变异体。在GH3细胞的染色质提取物，GH3细胞的催乳素阴性GC细胞变体和正常大鼠垂体组织中检测到NP44和NP48的水平相似。在大鼠肝脏和大鼠C6神经胶质细胞的染色质提取物中发现这两种蛋白的含量要低得多。 NP44和NP48表现出DNA序列特异性，这是由于它们与催乳激素基因的上游侧翼区域牢固结合，而与质粒DNA，大鼠催乳素或生长激素cDNA或其他两个大鼠基因的上游侧翼区域结合非常弱。通过分析大鼠催乳素基因组克隆的亚克隆，我们确定NP44和NP48结合至少两个位点，这些位点位于转录起始上游的0.4和2.0 kb之间（I区）和2.0到4.8 kb之间（II区）现场。这些发现是在NP44和NP48与催乳素5'侧翼区域的强结合与泌乳素基因的垂体特异性表达之间可能存在的功能关联的背景下进行讨论的。

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