Takata Y, Sagara Y, Kono A, Sekimizu K, Horiuchi T
Chemotherapy Division of Research Institute, National Kyushu Cancer Center, Fukuoka, Japan.
Biol Pharm Bull. 1993 Dec;16(12):1200-6. doi: 10.1248/bpb.16.1200.
The gene structure of bovine adrenodoxin reductase, a component of the mitochondrial steroid hydroxylating system in the adrenal cortex, was determined. When we screened a bovine genomic DNA library, using bovine adrenodoxin reductase cDNA as a probe, we isolated 10 genomic clones covering a continuous 30 kilobase (kb) region of bovine chromosomal DNA in which the adrenodoxin reductase gene spanned approximately 12 kb. The adrenodoxin reductase gene consisted of 12 exons and all the donor and acceptor sites of the exon/intron junction followed the GT/AG rule. The transcription initiation site was found to be 79 bases upstream of the translation initiation site by primer extension analysis and putative CAAT and GC boxes, but no typical TATA box, were present in the 5'-flanking region. The 5'-flanking region of this gene showed several features characteristic of promoters of house-keeping genes. The CCAGGG sequence present in the 5'-flanking region of genes for adrenodoxin and the steroid hydroxylating cytochromes P-450 in the adrenal cortex was present 13 bases upstream from the transcription initiation site. We analyzed the promoter activity of the 5'-upstream region of this gene, using as a reporter the chloramphenicol acetyltransferase gene. The region between position -757 to -376 was proved necessary for expression of this gene in mouse Y-1 tumor cells. Adrenocorticotropin did not enhance the expression of this bovine gene in the cells.
确定了牛肾上腺皮质线粒体类固醇羟化系统的一个组成部分——牛肾上腺皮质铁氧化还原蛋白还原酶的基因结构。当我们使用牛肾上腺皮质铁氧化还原蛋白还原酶cDNA作为探针筛选牛基因组DNA文库时，分离出了10个基因组克隆，它们覆盖了牛染色体DNA上一个连续的30千碱基（kb）区域，其中肾上腺皮质铁氧化还原蛋白还原酶基因跨度约为12 kb。肾上腺皮质铁氧化还原蛋白还原酶基因由12个外显子组成，外显子/内含子连接处的所有供体和受体位点均遵循GT/AG规则。通过引物延伸分析发现转录起始位点在翻译起始位点上游79个碱基处，在5'侧翼区域存在推测的CAAT盒和GC盒，但没有典型的TATA盒。该基因的5'侧翼区域表现出一些管家基因启动子的特征。肾上腺皮质铁氧化还原蛋白和类固醇羟化细胞色素P - 450基因的5'侧翼区域中存在的CCAGGG序列，位于转录起始位点上游13个碱基处。我们使用氯霉素乙酰转移酶基因作为报告基因，分析了该基因5'上游区域的启动子活性。结果证明，-757至-376位之间的区域对于该基因在小鼠Y - 1肿瘤细胞中的表达是必需的。促肾上腺皮质激素并未增强该牛基因在细胞中的表达。
