Molecular cloning of a novel transcriptional repressor protein of the rat type 1 vasoactive intestinal peptide receptor gene.

This study demonstrates that the transcriptional repressor sequence of the rat vasoactive intestinal peptide receptor (VIPR) gene constitutes a 42-base pair core element that is the binding site for a nuclear protein. We showed that this element was able to confer transcriptional repression to a heterologous promoter and that deletion or point mutations within this element resulted in loss of transcriptional repression. Southwestern blot analysis indicated that the VIPR repressor element interacts specifically with a nuclear protein of about 72 kDa. By screening a rat lung expression library coupled with rapid amplification of cDNA ends polymerase chain reactions, we isolated a cDNA clone (designated as VIPR-RP) that contains an open reading frame of 656 amino acids. VIPR-RP is 78% identical to a previously characterized protein, differentiation-specific element-binding protein, which is a member of a family of proteins including components of the DNA replication factor C complex. However, VIPR-RP cDNA encodes for a much smaller protein than differentiation-specific element-binding protein because of a frameshift. VIPR-RP mRNA is expressed in multiple tissues, including lung, liver, brain, heart, kidney, spleen, and testis. VIPR-RP protein specifically interacts with the VIPR repressor element as demonstrated by gel shift assays. Transfection of VIP-RP expression vector into Cos cells resulted in transcriptional repression of a reporter construct containing multiple copies of the VIPR repressor element. These results indicate that VIPR-RP is a novel transcriptional repressor protein that regulates VIPR expression.