Identification and characterization of the cell type-specific and developmentally regulated alpha7 integrin gene promoter

Abstract

Expression of alpha7 is mainly confined to skeletal and cardiac muscle in which it appears to be the major laminin-binding integrin. When myoblasts differentiate to myotubes, alpha7 mRNA and protein expression is up-regulated. To explore the mechanisms involved in the tissue-specific and developmentally regulated expression of alpha7, we isolated and characterized a genomic clone containing approximately 2.8 kilobase pairs (kb) of the 5'-flanking region of the murine alpha7 gene. The 5'-flanking region lacks both TATA and CCAAT boxes but contains five putative Sp1 binding sites located in a CpG island. Two transcription start sites, located near an initiator-like sequence, are 176 and 170 base pairs upstream of the translation start site. There are numerous binding sites for developmental and cell type-specific transcription factors, including AP-1, AP-2, GATA, and several AT-rich sites. There are also eight consensus E-boxes that bind the basic helix-loop-helix family of muscle-specific transcription factors. The approximately 2.8-kb 5'-flanking region was an active promoter in C2C12 skeletal myoblasts and exhibited increased expression upon conversion to myotubes but was inactive in HtLM2 cells, a mouse breast carcinoma epithelial cell line that does not express alpha7. Deletion analysis identified both positive and negative regulatory elements within the approximately 2.8-kb fragment. In 10T1/2 fibroblasts the approximately 2.8-kb alpha7 promoter was trans-activated by the myogenic basic helix-loop-helix proteins myogenin and MyoD but not by MRF4 and myf5. These results suggest that muscle-specific transcription factors play a role in regulating the cell-type expression of the alpha7 gene during development.