Glucocorticoid receptor binding sites in the promoter region of milk protein genes

The action of glucocorticoids on the induction of the two milk protein genes beta-casein and whey acidic protein (WAP) is delayed and appears to be indirect. The response requires a co-stimulation of cells with prolactin, is restricted to mammary epithelial cells and is dependent on cis-acting sequences localized in the promoter region of the two genes. We have searched for glucocorticoid receptor (GR) binding sites in these hormone response regions. In vitro DNaseI footprinting experiments were performed with a purified preparation of the GR from rat liver. The sequences between -231 and -7 and between -250 and -79 of mouse WAP and rat beta-casein gene promoter, respectively, were found to contain multiple sites which were protected from the attack of DNaseI by the GR preparation. The sites comprise sequence motifs related to the delayed secondary glucocorticoid response elements described (Chan et al., J. Biol. Chem. 266, 22,634-22,644, 1991), which represent half sites of classical GR binding sites. In the WAP gene promoter, the motifs are arranged head to tail with irregular spacing. The GR binding sites are in close proximity or even overlap with the bindings sites for candidate factors involved in mammary cell specific gene expression. The results suggest a direct co-operation between GR and mammary cell specific transcription factors in mediating the expression of milk protein genes.