The synthesis of protein requires the availability of specific AA and a large supply of energy in bovine mammary epithelial cells (BMEC). Whether an interaction exists between Lys/Met ratio and glucose level on milk protein synthesis and its potential regulatory mechanism is unclear. We investigated the effects of different Lys/Met ratios and glucose levels on casein synthesis-related gene expression in BMEC to elucidate the underlying molecular mechanisms. Primary BMEC were subjected to 4 treatments for 36 h, arranged in a 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 glucose levels of 17.5 mM (high glucose level) and 2.5 mM (low glucose level). No interactions between Lys/Met ratio and glucose level on cell viability, cell cycle progression, mRNA, or protein expression levels were found. High glucose level increased cell proliferation and promoted cell cycle transition from intermediate phase (G1 phase) to synthesis (S phase) by approximately 50%, whereas Lys/Met ratio had no effect. Both mRNA and protein abundance of αS1-casein and β-casein were positively affected by LM3.0, whereas a high glucose level increased protein abundance of αS1-casein and β-casein and increased gene expression of CSN1S1 but not of CSN2. Furthermore, high glucose increased the mRNA abundance of ELF5 and decreased that of GLUT8, enhanced protein expression of total and phosphorylated mechanistic target of rapamycin (mTOR), and decreased phosphorylated AMP-activated protein kinase (AMPK) levels. Treatment LM3.0 had a stimulatory effect on total and phosphorylated mTOR but did not affect AMPK phosphorylation. The mRNA levels of JAK2, ELF5, and RPS6KB1 were upregulated and mRNA levels of EIF4EBP1 were downregulated with LM3.0 compared with LM2.3. Our results indicate that casein synthesis was regulated by Lys/Met ratio via JAK2/ELF5, mTOR, and its downstream RPS6KB1 and EIF4EBP1 signaling. In contrast, glucose regulated casein synthesis through promoting cell proliferation, accelerating cell cycle progression, and activating the ELF5 and AMPK/mTOR signaling pathways. Within the range of substrate levels in the present study, a change in Lys/Met ratio had a stronger effect on abundance of αS1-casein and β-casein than a change in glucose level.
Tag: mineral content
Long-term accumulation of macro- and secondary elements in subtropical treatment wetlands
The Everglades Stormwater Treatment Areas (STAs) are a complex of large constructed wetlands that are an integral component of the State and Federal efforts to restore the Everglades ecosystem. The overall objective of this study was to determine the accumulation rates of macro-elements including carbon (C), nitrogen (N), phosphorus (P), sulfur (S), and associated secondary elements including calcium (Ca), magnesium (Mg), aluminum (Al), and iron (Fe) in two Everglades STAs over their periods of operation. The study was conducted in STA-2 with parallel flow-ways consisting of emergent aquatic vegetation (EAV) and submerged aquatic vegetation (SAV) and the Western flow-way of STA-3/4 with EAV and SAV cells operated in series. Elemental accumulation rates were determined in the unconsolidated surficial sediments (floc) and recently accreted soil (RAS) that have accumulated on top of the antecedent soil over the 14- and 10-yr periods of operation for STA-2 and STA-3/4, respectively. Flow-ways with SAV were more efficient than EAV in accreting mineral matter, resulting in increased bulk density and higher accumulation rates of elements. Average C accumulation in the floc and RAS of SAV flow-ways was 320 g·m−2·yr−1 with approximately equal proportions of inorganic and organic C, while in the EAV flow-ways accumulation rates of C ranged from 116 to 147 g·m−2·yr−1 with mostly organic C. Phosphorus accumulation rates were approximately 2–3 times higher in SAV than in EAV flow-ways. Differences in accumulation of elements between SAV and EAV were largest for Ca with 17–42 times more Ca in SAV than EAV systems. This suggests that in the SAV systems, possible occlusion of macro-elements and metals during CaCO3 precipitation facilitated accretion of material with high mineral content. In EAV, biomass turnover and associated biotic processes regulated organic matter accumulation rates. The spatial accumulation patterns of P, C, and N in the EAV areas of STA-2 and STA-3/4 were similar to those observed in the EAV areas of the natural wetlands in Water Conservation Areas, suggesting that constructed wetland systems function similarly to natural wetlands dominated by EAV areas in retaining and storing macro- and secondary elements.