Short communication: Relationship between lysine/methionine ratios and glucose levels and their effects on casein synthesis via activation of the mechanistic target of rapamycin signaling pathway in bovine mammary epithelial cells

摘要

The synthesis of protein requires the availabilityof specific AA and a large supply of energy in bovinemammary epithelial cells (BMEC). Whether an interactionexists between Lys/Met ratio and glucose levelon milk protein synthesis and its potential regulatorymechanism is unclear. We investigated the effects ofdifferent Lys/Met ratios and glucose levels on caseinsynthesis-related gene expression in BMEC to elucidatethe underlying molecular mechanisms. Primary BMECwere subjected to 4 treatments for 36 h, arranged ina 2 × 2 factorial design with Lys/Met ratios of 3:1(1.2:0.4 mM, LM3.0; total AA = 8.24 mM) and 2.3:1(1.4:0.6 mM, LM2.3; total AA = 8.64 mM) and glucoselevels of 17.5 mM (high glucose level) and 2.5 mM (lowglucose level). No interactions between Lys/Met ratioand glucose level on cell viability, cell cycle progression,mRNA, or protein expression levels were found.High glucose level increased cell proliferation and promotedcell cycle transition from intermediate phase (G1phase) to synthesis (S phase) by approximately 50%,whereas Lys/Met ratio had no effect. Both mRNA andprotein abundance of αS1-casein and β-casein were positivelyaffected by LM3.0, whereas a high glucose levelincreased protein abundance of αS1-casein and β-caseinand increased gene expression of CSN1S1 but not ofCSN2. Furthermore, high glucose increased the mRNAabundance of ELF5 and decreased that of GLUT8,enhanced protein expression of total and phosphorylatedmechanistic target of rapamycin (mTOR), anddecreased phosphorylated AMP-activated protein kinase(AMPK) levels. Treatment LM3.0 had a stimulatoryeffect on total and phosphorylated mTOR but didnot affect AMPK phosphorylation. The mRNA levelsof JAK2, ELF5, and RPS6KB1 were upregulated andmRNA levels of EIF4EBP1 were downregulated withLM3.0 compared with LM2.3. Our results indicate thatcasein synthesis was regulated by Lys/Met ratio viaJAK2/ELF5, mTOR, and its downstream RPS6KB1and EIF4EBP1 signaling. In contrast, glucose regulatedcasein synthesis through promoting cell proliferation,accelerating cell cycle progression, and activating theELF5 and AMPK/mTOR signaling pathways. Withinthe range of substrate levels in the present study,a change in Lys/Met ratio had a stronger effect onabundance of αS1-casein and β-casein than a change inglucose level.