Dipeptide (Methionyl-Methionine) Transport and Its Effect on β-Casein Synthesis in Bovine Mammary Epithelial Cells

作者：Wang, Caihong; Zhao, Fengqi; Liu, Jianxin; Liu, Hongyun^*

来源：Cellular Physiology and Biochemistry, 2018, 49(2): 479-488.

DOI：10.1159/000492987

• Bovine mammary epithelial cell
• beta-casein synthesis
• Dipeptide
• Methionyl-methionine
• mTOR signaling pathway
• Peptide transporter 2

Background/Aims: The aim of this study was to investigate the transport properties and utilization of methionyl-methionine dipeptide (Met-Met) in beta-casein (beta-CN) synthesis in bovine mammary epithelial cells (BMECs). Methods: The transport properties were studied for the effects of time, pH, concentration, temperature and inhibitors using Met-Met-FITC in BMECs. BMECs were treated with different concentrations of Met-Met (0, 20, 40, 80, 120 and 160 mu g/ml). In several experiments, the cells were treated with Janus kinase 2 (JAK2) inhibitor (tyrphostin AG-490, 50 mu M) and mammalian target of rapamycin (mTOR) inhibitor (rapamycin, 100 ng/ml). Results: The uptake of Met-Met-FITC by BMECs was rapid during the first fifteen minutes and became saturated after 15 minutes. The transport of Met-Met-FITC in BMECs exhibited a Michaelis constant of 52.4 mu M and maximum transport velocity of 14.8 pmol/min/mg protein. The uptake of Met-Met-FITC in BMECs was pH-dependent, peaked at pH 6.5 and was significantly inhibited by other peptides, including Met-Lys, Lys-Lys, Gly-Met, Gly-Leu and Met-Leu. Knocking down the peptide transporter 2 (PepT2) with small interference RNA markedly decreased Met-Met-FITC uptake. Met-Met concentration-dependently increased the PepT2 expression and beta-CN synthesis in BMECs with an optimal concentration of 80 mu g/ml. At 80 mu g/ml, Met-Met also enhanced the cell viability and cyclin D1 expression and promoted cell cycle transition from G1 phase to S phase. In addition, 80 mu g/ml Met-Met increased the mRNA abundance of JAK2 and signal transducer and activator of transcription 5 (STAT5) and enhanced the phosphorylation of JAK2, STAT5, mTOR, p70 ribosomal S6 kinase 1 and eukaryotic initiation factor 4E binding protein 1. The inhibition of JAK2 and mTOR significantly decreased Met-Met-induced increase in cell viability and beta-CN synthesis in BMECs. Conclusion: Our data elucidated the properties of peptide transporter and its effect on beta-CN synthesis in BMECs. Met-Met, taken up by PepT2, enhances cell proliferation and promotes beta-CN synthesis by activating JAK2-STAT5 and mTOR signaling pathways in BMECs.

• 出版日期2018
• 单位浙江大学