Prolactin regulation of beta-casein gene expression and of a cytosolic 120-kd protein in a cloned mouse mammary epithelial cell line.

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Abstract
In order to study the hormonal regulation of gene expression in mammary epithelial cells, we isolated a prolactin‐responsive cell clone, HC11, from the COMMA‐1D mouse mammary epithelial cell line. Clone HC11 was selected as a unique example of a cloned mouse mammary epithelial cell which has no requirement for complex, exogenously added, extracellular matrix or co‐cultivation with other cell types for the prolactin‐dependent in vitro induction of the endogenous beta‐casein gene by lactogenic hormones. Induction of beta‐casein mRNA is rapid and was detected 3 h after hormone stimulation. A prolactin‐dependent increase in the rate of transcription of the beta‐casein gene was shown in an in vitro nuclear transcription assay. beta‐Casein protein was detected in an immunoblot assay after 24 h, and further accumulated during 5 days of hormone treatment. To identify low‐abundance proteins induced directly after prolactin stimulation, mRNA was accumulated during 5 h of stimulation of HC11 cells with prolactin in the presence of cycloheximide. Following cycloheximide removal, the mRNA was translated into protein during a 60‐min [35S]methionine pulse and the proteins were resolved by DEAE ion exchange HPLC and SDS‐PAGE. A strong induction of a 120‐kd cytosolic protein was detected which was maximally expressed within 6 h of hormone stimulation.