Identification of two regulatory binding sites which confer myotube specific expression of the mono-ADP-ribosyltransferase ART1 gene
Identification of two regulatory binding sites which confer myotube specific expression of the mono-ADP-ribosyltransferase ART1 gene
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DOI：
10.1186/1471-2199-9-91
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发表时间：
2008-10-21
影响因子：
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通讯作者：
Hauschildt, Sunna
中科院分区：
文献类型：
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作者：
Friedrich, Maik;Boehlig, Levin;Hauschildt, Sunna
背景资料：单ADP核糖基转移酶（ART）1属于哺乳动物胞外酶家族，催化ADP核糖从NAD（+）转移到靶蛋白。ART 1主要在骨骼肌和心肌中表达。其ADP-核糖基化α 7-整联蛋白，α 7-整联蛋白与β 1-整联蛋白一起形成二聚体并结合层粘连蛋白，层粘连蛋白是参与细胞粘附的细胞外基质的蛋白质。这种翻译后修饰导致层粘连蛋白结合亲和力增加。结果：使用C2 C1/2和C3 H-10 T 1/2细胞作为模型的肌生成，我们发现，ART 1的表达仅限于肌管形成。我们确定了一个片段跨越基因上游1.3 kb的转录起始位点作为ART 1基因的功能性启动子。该区域包含一个E盒和一个富含A/T的元件，这两个保守的转录因子结合位点存在于大多数骨骼肌特异性基因的启动子中。突变的DNA序列的E盒或A/T-丰富的元素导致ART 1启动子诱导几乎完全丧失，表明这些网站的转录因子结合的合作作用。凝胶迁移率变动分析进行了核提取物从C2 C1/2和C3 H-10 T 1/2细胞显示肌细胞生成素的E盒和MEF-2的A/T丰富的元素，结合被限制到C2 C1/2和C3 H-10 T 1/2 myotubes.Conclusion：在这里，我们描述的分子机制的ART 1基因表达的调控在骨骼肌细胞。ART 1 mRNA的分化依赖性上调是由肌细胞生成素与ART 1基因近端启动子区的E盒和MEF-2与富含A/T的元件的结合诱导的。因此，转录调控涉及的分子机制类似于那些用于激活肌肉特异性基因。
Background: Mono-ADP-ribosyltransferase (ART) 1 belongs to a family of mammalian ectoenzymes that catalyze the transfer of ADP-ribose from NAD(+) to a target protein. ART1 is predominantly expressed in skeletal and cardiac muscle. It ADP-ribosylates alpha 7-integrin which together with beta 1-integrin forms a dimer and binds to laminin, a protein of the extracellular matrix involved in cell adhesion. This posttranslational modification leads to an increased laminin binding affinity.Results: Using C2C12 and C3H-10T 1/2 cells as models of myogenesis, we found that ART1 expression was restricted to myotube formation. We identified a fragment spanning the gene 1.3 kb upstream of the transcriptional start site as the functional promoter of the ART1 gene. This region contains an E box and an A/T-rich element, two conserved binding sites for transcription factors found in the promoters of most skeletal muscle specific genes. Mutating the DNA consensus sequence of either the E box or the A/T-rich element resulted in a nearly complete loss of ART1 promoter inducibility, indicating a cooperative role of the transcription factors binding to those sites. Gel mobility shift analyses carried out with nuclear extracts from C2C12 and C3H-10T 1/2 cells revealed binding of myogenin to the E box and MEF-2 to the A/T-rich element, the binding being restricted to C2C12 and C3H-10T 1/2 myotubes.Conclusion: Here we describe the molecular mechanism underlying the regulation of the ART1 gene expression in skeletal muscle cells. The differentiation-dependent upregulation of ART1 mRNA is induced by the binding of myogenin to an E box and of MEF-2 to an A/T-rich element in the proximal promoter region of the ART1 gene. Thus the transcriptional regulation involves molecular mechanisms similar to those used to activate muscle-specific genes.