Controlled synchronization of prolactin/STAT5 and AKT1/mTOR in bovine mammary epithelial cells

The prolactin/STAT5 and AKT1/mTOR pathways play a key role in milk protein transcription and translation, respectively. However, the correlation between them in bovine mammary epithelial cells remains unclear. Here, mRNA and protein expression levels of AKT1, STAT5, and mTOR and the phosphorylation of these proteins were determined. Cell proliferation and viability were examined using the CASY-TT assay. Purified bovine mammary epithelial cells were cultured in differentiation media for different periods. The basic differentiation medium contained a lactogenic hormone cocktail of insulin (5 μg/mL), hydrocortisone (1 μg/mL), and prolactin (5 μg/mL). The cells cultured in this medium grew slowly and expressed higher levels of p-STAT5, p-AKT1, and p-mTOR (activated form) than those of control cells. Although the phosphorylation ratio was not increased, transcription and translation of these proteins were upregulated by the addition of insulin-like growth factor-1 or growth hormone, which further increased β-casein mRNA expression. Furthermore, the three proteins were upregulated or downregulated synchronously, even after RNA interference silencing of either Stat5 or Akt1. These findings indicate that a few hormones and other factors play lactogenic and galactogenic roles by promoting two key lactogenic signaling associated with milk protein expression. We provide evidence of prolactin/STAT5 and AKT1/mTOR synchronization, establishing a direct correlation between transcription regulation and translation regulation of milk protein in bovine mammary epithelial cells.