The rat Mist1 gene: structure and promoter characterization

Transcription factors of the basic Helix-Loop-Helix (bHLH) protein family p lay key roles in several developmental processes. Mist1 belongs to this gro up of proteins and shares several properties with the other family members. For example, Mist1 is capable of dimerization with the ubiquitously expres sed E2A bHLH proteins and exhibits a strong DNA-binding activity to the cor e E-box sequence. Using in-situ hybridization and Northern blot hybridizati on, Mist1 mRNA has been detected in a variety of embryonic and adult rodent tissues. To understand the molecular mechanisms involved in the expression of the gene, we have cloned the rat Mist1 gene and analyzed 2.5 kb of its 5' flanking region. The Mist1 gene spans over 5 kilobases and is composed o f two exons separated by a unique intron. The entire coding region is local ized in the second exon. Sequence analysis of the promoter region indicated an absence of TATA-box or CAAT-box sequence, but several consensus Sp1-bin ding sites were present near the transcription start site. Deletion analysi s of the promoter region identified a 272 bp proximal fragment to be suffic ient to drive expression of a reporter gene in N1H3T3 fibroblasts. Subseque nt deletion of potential Sp1 sites results in a marked decrease in promoter activity. Electrophoretic mobility shift assays revealed that Sp1 binds to two different regions in the proximal promoter, a typical Sp1 site located at (-38; -33) and a G/C-rich region between (-67; -62). These data suggest that the basal expression of this TATA-less gene might be driven by genera l transcription factors, such as Sp1. (C) 2000 Elsevier Science B.V. All ri ghts reserved.