Human Mullerian-inhibiting substance promoter contains a functional TFII-I-binding initiator

Mullerian-inhibiting substance (MIS) plays an essential role in mammalian m ale sexual development; thus, it is important to determine how the tightly regulated expression of the MIS gene is transcriptionally controlled. Trans cription of eukaryotic genes is dependent on regulatory elements in the enh ancer and one or both distinct elements in the core promoter: the TATA box, and the initiator (Inr) element. Because the human MIS gene does not conta in a consensus TATA and has not been reported to contain an Inr element, we hypothesized that the initiator region of the core promoter was essential for promoter activity. Transient transfection assays were conducted using a n immortalized Embryonic Day 14.5 male rat urogenital ridge cell line (CH34 ) that expresses low levels of MIS. These studies revealed that promoter ac tivity is dependent on the region around the start site (-6 to +10) but not on the nonconsensus TATA region. Electrophoretic mobility shift assays dem onstrated that the human MIS initiator sequence forms a specific DNA-protei n complex with CH34 cell nuclear extract, HeLa cell nuclear extract, and pu rified TFII-I. This complex could be blocked or supershifted by the additio n of antibodies directed against TFII-I. These data suggest that the human MIS gene contains a functional initiator that is specifically recognized by TFII-I.