Kemp D M, Habener J F
Laboratory of Molecular Endocrinology, Massachusetts General Hospital, Howard Hughes Medical Institute, Harvard Medical School, Boston, Massachusetts 02114, USA.
Endocrinology. 2001 Mar;142(3):1179-87. doi: 10.1210/endo.142.3.8026.
The insulin gene promoter contains many transcriptional response elements that predispose the gene to a wide range of regulatory signals. Glucagon-like peptide 1 (GLP-1) stimulates insulin gene transcription by intracellular second messenger cascades leading to direct transcription factor activation or to the up-regulation of insulin promoter specific transcription factors. In these studies, we have identified a novel regulatory signaling mechanism acting on the rat insulin 1 promoter (rINS1) in the INS-1 beta-cell line. In the presence of stimulatory concentrations of GLP-1 (0.1--100 nM) on rINS1 activity, inhibition of p38 mitogen-activated protein kinase (p38 MAPK) using SB 203580 resulted in a marked increase in promoter activity (maximum 3-fold) over GLP-1 alone, as determined by rINS1 promoter-luciferase reporter gene expression. This effect was revealed to be mediated via the cAMP response element (CRE) of rINS1, because site directed mutagenesis of the CRE motif in rINS1 abolished the increased response to SB 203580. Furthermore, inhibition of p38 MAPK uncovered a similar, more pronounced, response in the expression of a generic CRE promoter driven reporter gene. Time course dose-response studies indicate that the p38 MAPK induced inhibitory response may involve expression of immediate early genes (IEGs); maximum repression of rINS1 activity occurred after 4 h of treatment, comparable with regulatory responses by IEGs. In conclusion, these results demonstrate a novel signaling mechanism whereby p38 MAPK represses rINS1 promoter activity in response to GLP-1, suggesting the involvement of a robust regulatory control by p38 MAPK in insulin gene expression. The relevance of this mechanism may be most apparent during periods of cellular stress in which p38 MAPK activity is stimulated. In this regard, reduced insulin expression levels caused by chronic hyperglycemia (glucotoxicity) and/or hyperlipidemia (lipotoxicity) may be a direct consequence of this mechanism.
胰岛素基因启动子包含许多转录反应元件，使该基因易于受到多种调节信号的影响。胰高血糖素样肽1（GLP-1）通过细胞内第二信使级联反应刺激胰岛素基因转录，导致直接激活转录因子或上调胰岛素启动子特异性转录因子。在这些研究中，我们在INS-1β细胞系中鉴定出一种作用于大鼠胰岛素1启动子（rINS1）的新型调节信号机制。当存在刺激浓度的GLP-1（0.1 - 100 nM）时，使用SB 203580抑制p38丝裂原活化蛋白激酶（p38 MAPK），与单独使用GLP-1相比，rINS1启动子荧光素酶报告基因表达显示启动子活性显著增加（最大3倍）。这种效应被证明是通过rINS1的cAMP反应元件（CRE）介导的，因为rINS1中CRE基序的定点诱变消除了对SB 203580增加的反应。此外，抑制p38 MAPK在通用CRE启动子驱动的报告基因表达中发现了类似的、更明显的反应。时间进程剂量反应研究表明，p38 MAPK诱导的抑制反应可能涉及即刻早期基因（IEGs）的表达；处理4小时后rINS1活性出现最大抑制，这与IEGs的调节反应相当。总之，这些结果证明了一种新型信号机制，即p38 MAPK响应GLP-1抑制rINS1启动子活性，表明p38 MAPK在胰岛素基因表达中参与了强大的调节控制。在细胞应激期间p38 MAPK活性受到刺激时，这种机制的相关性可能最为明显。在这方面，慢性高血糖（糖毒性）和/或高血脂（脂毒性）导致的胰岛素表达水平降低可能是这种机制的直接后果。
