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

Abstract

Background/Aims: The aim of this study was to investigate the transport properties and utilization of methionyl-methionine dipeptide (Met-Met) in β-casein (β-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 µg/ml). In several experiments, the cells were treated with Janus kinase 2 (JAK2) inhibitor (tyrphostin AG-490, 50 µM) 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 µM 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 β-CN synthesis in BMECs with an optimal concentration of 80 µg/ml. At 80 µg/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 µg/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 β-CN synthesis in BMECs. Conclusion: Our data elucidated the properties of peptide transporter and its effect on β-CN synthesis in BMECs. Met-Met, taken up by PepT2, enhances cell proliferation and promotes β-CN synthesis by activating JAK2-STAT5 and mTOR signaling pathways in BMECs.

Effects of methionine partially replaced by methionyl‐methionine dipeptide on intestinal function in methionine‐deficient pregnant mice

“…In addition, our results further suggest that altered TJ proteins from can enhance the cell cycle transition from G1 phase to S phase in bovine mammary epithelial cells (Wang, Zhao, Liu, & Liu, 2018).…”

“…Met or Met‐Met supplementation may have different effects on one‐carbon metabolism pathway in the liver, the major organ for Met metabolism (Bauchart‐Thevret, Stoll, Chacko, et al, ). Another possibility may be associated with the altered levels of cell apoptosis in the intestine in two groups as we found that Met‐Met supplementation can enhance the cell cycle transition from G1 phase to S phase in bovine mammary epithelial cells (Wang, Zhao, Liu, & Liu, ). Apoptosis is critical for intestinal epithelial turnover and homeostasis, but excessive apoptosis can lead to intestinal dysfunction (Günther, Neumann, Neurath, & Becker, ).…”

Methionine Partially Replaced by Methionyl-Methionine Dipeptide Improves Reproductive Performance over Methionine Alone in Methionine-Deficient Mice

“…It was first found in 1996 that small peptide-containing Met could serve as a source of free Met for milk protein synthesis, and the efficiency of some Met dipeptides was much higher than the equivalent FAA in mouse mammary gland tissue explants [ 9 ]. Recently, we identified that small peptides containing Met, Lys, or Phe could promote αS1-casein and β-casein synthesis in bovine mammary gland explants [ 10 , 11 , 12 , 13 ]. Apart from its role as protein synthesis substrates, small peptides may also function as signaling molecules to promote cell cycle transition and AA absorption [ 12 , 13 ].…”

“…Recently, we identified that small peptides containing Met, Lys, or Phe could promote αS1-casein and β-casein synthesis in bovine mammary gland explants [ 10 , 11 , 12 , 13 ]. Apart from its role as protein synthesis substrates, small peptides may also function as signaling molecules to promote cell cycle transition and AA absorption [ 12 , 13 ]. Although the role of dipeptides in milk protein synthesis in vitro is clearly established, little is known about its role in fetal development.…”

Identification of octopus peptide and its promotion of β‐casein synthesis in a mouse mammary epithelial cell line

“…And several studies show that peptides could be utilized as an amino nitrogen by the mammary gland for milk protein production. Researches elucidated that Met‐containing di‐ to octapeptides promoted cell proliferation and increased β‐casein production by activating JAK2‐STAT5 and mTOR signaling pathways in mammary epithelial cells (Guo, Dan, Cao, Khas‐Erdene, & C., 2017; Pan & Webb, 1998; Wang, Zhao, Liu, & Liu, 2018). Studies suggest that Lys is a primary limiting amino acid for mammary gland development and simulates protein synthesis by activating the mTOR pathway (Lin et al., 2018).…”