Induction of utrophin gene expression by heregulin in skeletal muscle cells: Role of the N-box motif and GA binding protein
Induction of utrophin gene expression by heregulin in skeletal muscle cells: Role of the N-box motif and GA binding protein
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DOI：
10.1073/pnas.96.6.3223
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发表时间：
1999-03-16
影响因子：
11.1
通讯作者：
Jasmin, BJ
中科院分区：
文献类型：
--
作者：
Gramolini, AO;Angus, LM;Jasmin, BJ
神经源性因子agrin对肌肉中肌营养因子ARIA/heregulin基因表达的调节似乎涉及到营养因子ARIA/heregulin。在这里，我们发现heregulin处理小鼠和人类培养的肌管导致肌营养因子mRNA水平增加约2.5倍。瞬时转染肌营养因子启动子报告基因的实验表明，这种增加是由于肌营养因子基因转录的增强。在烟碱乙酰胆碱受体delta和epsilon亚基基因的情况下，先前报道过heregulin通过一个保守的启动子元件N-box刺激转录，该元件与多聚体ets相关转录因子GA结合蛋白（GABP）结合。因此，在肌营养蛋白基因启动子中单个N-box基序的位点定向突变消除了对heregulin的转录反应。此外，heregulin或培养肌管中两个GABP亚基的过表达导致N-box依赖性的utrophin启动子活性增加。在体内，直接基因转移到肌肉中证实了heregulin调节肌营养因子基因的表达。最后，用肌肉提取物进行的电泳迁移位移分析和超位移实验表明，肌营养蛋白启动子的N-box与GABP结合，这些发现表明，heregulin通过N-box motif和GABP在神经肌肉连接处表达的基因中对亚突触的转录激活是保守的，因为肌营养蛋白可以在功能上弥补肌营养不良蛋白的缺乏。阐明调节肌营养因子基因转录的分子机制可能最终导致基于肌营养不良患者整个肌纤维中肌营养因子表达的治疗。
The modulation of utrophin gene expression in muscle by the nerve-derived factor agrin plausibly involves the trophic factor ARIA/heregulin, Here we show that heregulin treatment of mouse and human cultured myotubes caused a approximate to 2.5-fold increase in utrophin mRNA levels. Transient transfection experiments with utrophin promoter-reporter gene constructs showed that this increase resulted from an enhanced transcription of the utrophin gene. In the case of the nicotinic acetylcholine receptor delta and epsilon subunit genes, heregulin was previously reported to stimulate transcription via a conserved promoter element, the N-box, which binds the multimeric Ets-related transcription factor GA binding protein (GABP). Accordingly, site-directed mutagenesis of a single N-box motif in the utrophin gene promoter abolished the transcriptional response to heregulin, In addition, overexpression of heregulin, or of the two GABP subunits in cultured myotubes, caused an N-box-dependent increase of the utrophin promoter activity. In vivo, direct gene transfer into muscle confirmed that heregulin regulates utrophin gene expression. Finally, electrophoretic mobility shift assays and supershift experiments performed with muscle extracts revealed that the N-box of the utrophin promoter binds GABP, These findings suggest that the subsynaptic activation of transcription by heregulin via the N-box motif and GABP are conserved among genes expressed at the neuromuscular junction, Because utrophin can functionally compensate for the lack of dystrophin, the elucidation of the molecular mechanisms regulating utrophin gene transcription may ultimately lead to therapies based on utrophin expression throughout the muscle fibers of Duchenne muscular dystrophy patients.