Friedrich Maik, Böhlig Levin, Kirschner Ralf D, Engeland Kurt, Hauschildt Sunna
Institute of Biology II, Dept, of Immunobiology, University of Leipzig, Talstrasse 33, D-04103 Leipzig, Germany.
BMC Mol Biol. 2008 Oct 21;9:91. doi: 10.1186/1471-2199-9-91.
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 alpha7-integrin which together with beta1-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.
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.
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.
单（ADP-核糖）基转移酶（ART）1属于哺乳动物胞外酶家族，可催化将ADP-核糖从NAD⁺转移至靶蛋白。ART1主要在骨骼肌和心肌中表达。它将α7整合素进行ADP-核糖基化，α7整合素与β1整合素一起形成二聚体，并与层粘连蛋白结合，层粘连蛋白是细胞外基质中参与细胞黏附的一种蛋白质。这种翻译后修饰导致层粘连蛋白结合亲和力增加。
使用C2C12和C3H-10T 1/2细胞作为肌生成模型，我们发现ART1的表达仅限于肌管形成。我们鉴定出一个跨越转录起始位点上游1.3 kb基因的片段作为ART1基因的功能启动子。该区域包含一个E盒和一个富含A/T的元件，这是在大多数骨骼肌特异性基因启动子中发现的两个保守转录因子结合位点。突变E盒或富含A/T元件的DNA共有序列会导致ART1启动子诱导性几乎完全丧失，表明结合到这些位点的转录因子具有协同作用。用C2C12和C3H-10T 1/2细胞的核提取物进行的凝胶迁移率变动分析显示，肌细胞生成素与E盒结合，MEF-2与富含A/T的元件结合，这种结合仅限于C2C12和C3H-10T 1/2肌管。
在此我们描述了骨骼肌细胞中ART1基因表达调控的分子机制。ART1 mRNA的分化依赖性上调是由肌细胞生成素与ART1基因近端启动子区域的E盒结合以及MEF-2与富含A/T的元件结合所诱导的。因此，转录调控涉及与用于激活肌肉特异性基因类似的分子机制。
