Regulation of cytochrome P-450b/e gene expression by a heme- and phenobarbitone-modulated transcription factor.
Regulation of cytochrome P-450b/e gene expression by a heme- and phenobarbitone-modulated transcription factor.
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血红素和苯巴比妥调节的转录因子对细胞色素 P-450b/e 基因表达的调节。
DOI：
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发表时间：
1989
影响因子：
11.1
通讯作者：
Govindarajan Padmanaban
中科院分区：
文献类型：
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作者：
P. Rangarajan;Govindarajan Padmanaban
克隆的细胞色素P-450B/e基因的DNA片段包含从-179位到第一外显子部分的上游区域，在冻融的大鼠肝细胞核中被忠实地转录。动物的苯巴比妥处理显著增加了这种转录，而动物的环己亚胺(蛋白质合成抑制剂)或CoCl2(血红素生物合成抑制剂)处理则阻断了这种增加。这张图片与报道的细胞色素P-450B/e在体内的mRNA水平和体外在这些条件下的连续转录速度非常相关。通过硝酸纤维滤膜结合、凝胶滞留、DNase I处理(“足迹”)和Western印迹分析，评估上游区域(从-179位)与核提取物结合的蛋白质。苯巴比妥处理显著增加了与上游区域的蛋白质结合，这一增加再次被放线菌亚胺或CoCl2处理所阻断。在体外将血红素添加到缺乏血红素的核和核提取物中，分别恢复了诱导的转录水平和与上游片段结合的蛋白质。因此，药物介导的合成和血红素调节的转录因子结合(S)似乎参与了细胞色素P-450B/e基因的转录激活，而85 kDa的蛋白可能是这方面的主要因素。
The cloned DNA fragment of the cytochrome P-450b/e gene containing the upstream region from position -179 through part of the first exon is faithfully transcribed in freeze-thawed rat liver nuclei. Phenobarbitone treatment of the animal strikingly increases this transcription, and the increase is blocked by cycloheximide (protein synthesis inhibitor) or CoCl2 (heme biosynthetic inhibitor) treatment of animals. This picture correlates very well with the reported cytochrome P-450b/e mRNA levels in vivo and run-on transcription rates in vitro under these conditions. The upstream region (from position -179) was assessed for protein binding with nuclear extracts by nitrocellulose filter binding, gel retardation, DNase I treatment ("footprinting"), and Western blot analysis. Phenobarbitone treatment dramatically increases protein binding to the upstream region, an increase once again blocked by cycloheximide or CoCl2 treatments. Addition of heme in vitro to heme-deficient nuclei and nuclear extracts restores the induced levels of transcription and protein binding to the upstream fragment, respectively. Thus, drug-mediated synthesis and heme-modulated binding of a transcription factor(s) appear involved in the transcriptional activation of the cytochrome P-450b/e genes, and an 85-kDa protein may be a major factor in this regard.