Identification and characterization of the minimal promoter of Mipu1: the role of GC boxes in the regulation of basal transcription

Mipu1, a novel gene encoding a KRAB/C2H2 zinc-finger protein, was first reported to be up-regulated in myocardial ischemia-reperfusion injury, functioning to protect cells against oxidative stress. To map the promoter region of the gene and to understand its regulation, we identified the transcription start site and revealed that the 1366-bp fragment upstream of the transcription start site possesses promoter activity. Deletion constructs of the 5 '-flanking region of Mipu1 lead to the identification of a minimal promoter, in which neither a TATA box nor a CAAT box was detected. Two GC boxes were found; however, they are the specific binding sites for Sp1-family transcription factors. Mutations in these GC boxes resulted in the total loss of Mipu1 minimal promoter activity. Finally, WP631, an Sp1-family-specific inhibitor, was found to decrease the promoter activity in a dose-dependent manner, indicating that the GC boxes are essential for the activity of the Mipu1 minimal promoter activity.