NEGATIVE REGULATORY ELEMENT IN THE MAMMARY SPECIFIC WHEY ACIDIC PROTEIN PROMOTER

Expression of the whey acidic protein (WAP) gene is tightly regulated in a tissue and developmental stage specific manner, in that the WAP g ene is exclusively expressed in the mammary gland during pregnancy and lactation. Using both deletion and competition analyses, evidence is provided for the existence of a negative regulatory element (NRE) in t he WAP promoter located between -413 and -93 with respect to the WAP t ranscriptional initiation site. This NRE dramatically decreases transc ription from linked heterologous promoter-reporter gene constructs. Th e activity of NRE requires WAP promoter sequences that are 230 bp apar t since subfragments of the NRE fail to inhibit transcription of adjoi ning reporter genes. Nuclear extracts from different cell types, in wh ich the WAP gene is not active, contain a protein or complex that spec ifically interacts with the entire NRE but not with subfragments of it . The contact points between this protein (NRE binding factor [NBF]) a nd the NRE element have been partially determined. Mutation of the imp licated nucleotides severely reduces the ability of NBF to bind, and s uch mutated promoter fragments fail to alleviate transcriptional repre ssion in competition experiments. This suggests that NBF binding to th e NRE is at least in part responsible for the negative regulation of t he WAP promoter. Since NBF is not detectable in the lactating mammary gland, where the WAP gene is expressed, we speculate that it may be a determinant of the expression spectrum of the WAP gene. (C) 1994 Wiley -Liss, Inc.