Prolactin regulatory element binding protein as a potential transcriptional factor for the insulin gene in response to glucose stimulation
Prolactin regulatory element binding protein as a potential transcriptional factor for the insulin gene in response to glucose stimulation
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DOI：
10.1007/s00125-006-0255-y
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发表时间：
2006-07-01
期刊：
影响因子：
8.2
通讯作者：
Ishida, T.
中科院分区：
文献类型：
--
作者：
Ohtsuka, S.;Murao, K.;Ishida, T.
目的/假设：催乳素调节元件结合蛋白(PREB)已被确定为调节大鼠垂体前叶催乳素启动子活性的因子。Preb不仅位于垂体前叶，也位于胰腺，但其在胰腺中的作用尚不清楚。因此，我们研究了PREB在胰岛素基因表达中的作用。材料和方法：采用荧光素酶报告基因分析和凝胶迁移率改变分析(EMSA)分析PREB对胰岛素基因转录的影响。在PREB表达或被敲除的细胞中，测定胰岛素的表达和分泌。结果：PreB主要定位于大鼠胰岛β细胞及其细胞系INS-1的胞核。INS-1细胞的核提取液含有可被PREB抗血清识别的物质。在EMSA研究中，这种核提取物还显示出被PREB抗血清超转移的胰岛素启动子结合活性。在INS-1细胞中，PREB和胰岛素启动子的共表达诱导了后者的活性。细胞中加入葡萄糖后，PREB的表达增加。胰岛素启动子的缺失分析表明，胰岛素启动子中的葡萄糖反应顺式元件A3介导了PREB的转录效应。此外，合成的PREB通过EMSA与A3元件结合，而该基序在胰岛素启动子中的突变体取消了PREB的作用。表达或下调PREB的细胞分别表现为胰岛素表达增加或减少。结论/解释：这些结果表明PREB可能参与了葡萄糖刺激下胰岛素基因转录和胰岛素分泌的调节。
Aims/hypothesis: Prolactin regulatory element binding (PREB) protein has been identified as a factor that regulates prolactin promoter activity in rat anterior pituitary. PREB is located not only in the anterior pituitary but also in pancreas; however its role in the pancreas is not known. We therefore examined the role of PREB in insulin gene expression. Materials and methods: To analyse the effects of PREB on insulin gene transcription, we employed the luciferase reporter gene assay and electrophoretic mobility shift assay (EMSA). In cells expressing or knocked down for PREB, insulin expression and secretion were determined. Results: PREB was located mainly in nuclei of rat pancreatic beta cells and its cell line, INS-1. A nuclear extract of INS-1 cells contained material that was recognised by PREB antiserum. This nuclear extract also showed insulin promoter binding activity that was super-shifted by PREB antiserum in EMSA studies. In the INS-1 cells, co-expression of PREB and the insulin promoter induced activity of the latter. The addition of glucose to the cells increased PREB expression. Deletional analysis of the insulin promoter showed that A3, a glucose-responsive cis-element in the insulin promoter, mediated the transcriptional effect of PREB. In addition, synthesised PREB bound the A3 element by EMSA, while a mutant of this motif in the insulin promoter abrogated the effect of PREB. Cells expressing or knocked down for PREB exhibited increased or decreased insulin expression, respectively. Conclusions/interpretation: These results demonstrate that PREB may contribute to the regulation of insulin gene transcription and insulin secretion in response to glucose stimulation.