CHARACTERIZATION OF AN UP-STREAM PROMOTER DIRECTING EXTRAPITUITARY EXPRESSION OF THE HUMAN PROLACTIN GENE
CHARACTERIZATION OF AN UP-STREAM PROMOTER DIRECTING EXTRAPITUITARY EXPRESSION OF THE HUMAN PROLACTIN GENE
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DOI：
10.1210/me.8.5.635
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发表时间：
1994-05-01
影响因子：
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通讯作者：
DAVIS, JRE
中科院分区：
文献类型：
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作者：
BERWAER, M;MARTIAL, JA;DAVIS, JRE
人 PRL 基因在垂体催乳素细胞外的蜕膜化子宫内膜和淋巴样细胞中表达，但此处 mRNA 包含来自附加非编码外显子 1a 的 5'-非翻译区。我们分离出含有人 PRL 外显子 1a 的基因组 DNA 克隆，其 5' 侧翼序列具有 2800 个碱基对 (bp)。测序定位于垂体起始位点上游的外显子 1a -5840 bp。为了研究其可疑的调节功能，将不同长度的 5' 侧翼区域与荧光素酶 (Luc) 报告基因连接起来。它们指导基因表达的能力已在转染研究中进行了分析。启动子序列的近端 1620 bp 引导 Luc 在 T 淋巴细胞 Jurkat 细胞系中表达，并且这不受近端 453 bp 的 5' 缺失的影响。然而，进一步对最近的 67 bp 进行 5' 删除后，该活性大大降低了 90%。外显子1a启动子在垂体GH细胞和HeLa细胞中失活；相比之下，外显子 1b 垂体启动子在 GH 细胞中活跃，但在 Jurkat 细胞中失活。 DNase-1 足迹研究以及进一步的 5' 和 3' 缺失分析确定了位于 -375/-212 bp 的增强子元件内的因子结合位点，该位点贡献了大约 50% 的启动子活性。
The human PRL gene is expressed outside pituitary lactotrophs in decidualized endometrium and lymphoid cells, but here the mRNA contains a 5'-untranslated region from an additional noncoding exon 1a. We have isolated a genomic DNA clone containing human PRL exon 1a with 2800 basepairs (bp) of 5'-flanking sequences. Sequencing locates exon 1a -5840 bp up-stream of the pituitary start site. To study its suspected regulatory function, various lengths of the 5'-flanking region were linked to the luciferase (Luc) reporter gene. Their ability to direct gene expression has been analyzed in transfection studies. The proximal 1620 bp of promoter sequence directed Luc expression in the T-lymphoid Jurkat cell line, and this was unaffected by 5'-deletion to the proximal 453 bp. However, further 5'-deletion to the most proximal 67 bp drastically reduced this activity by 90%. The exon 1a promoter was inactive in pituitary GH, cells and HeLa cells; in contrast, the exon 1b pituitary promoter, active in GH, cells, was inactive in Jurkat cells. DNase-1 footprinting studies and further 5'- and 3'-deletion analysis identified factor-binding sites within an enhancer element located at -375/-212 bp, which contributed approximately 50% of the promoter activity.