Characterization of the promoter for the mouse α3 integrin gene

The a3b1 integrin is an adhesion receptor for extracellular matrix proteins including isoforms of laminin, and the changes of its expression level in various cancer cells are thought to cause their malignant phenotypes. We have cloned an approximately 4 kb DNA fragment of the 5¢flanking region of the murine a3 integrin gene and analyzed its promoter activity. Transfection of MKN1 gastric carcinoma cells with serially truncated segments of the 5¢-flanking region linked to a luciferase gene indicated that a 537-bp SalI/SacI fragment upstream of exon 1 was sufficient to promote high level gene expression. By 5¢-rapid amplification of cDNA ends (5¢-RACE) using a cap site-labeled cDNA library, we determined one major and one minor transcription start sites in this region. The murine a 3i ntegrin gene was found to contain a CCAAT box, but to lack a TATA box. Luciferase assay following transfection with a series of deletion constructs of the SalI/SacI fragment revealed that the sequence between positions )260 and )119 bp (relative to the major transcription start site) is required for efficient transcription in gastric carcinoma cells. The sequence analysis of this segment showed the presence of several consensus sequences for transcription factors including Ets, GATA and MyoD/E-box binding factors. The introduction of mutation in one of the Ets-binding sequences greatly decreased its promoter activity, suggesting that the transcription of the a3 integrin gene in these cells is regulated by the Ets-family of transcription factors.