Effects of Lysine on Gene Expressions and Protein Phosphorylation Involved in Milk Protein Synthesis in Bovine Mammary Epithelial Cells

CHEN Lu, ZHAO Yanli, GUO Xiaoyu, SHI Binlin, YAN Sumei

doi:10.3969/j.issn.1006-267x.2018.08.032

2018 , Vol. 30 >Issue 8: 3142 - 3150

DOI: https://doi.org/10.3969/j.issn.1006-267x.2018.08.032

Molecular Nutrition

Effects of Lysine on Gene Expressions and Protein Phosphorylation Involved in Milk Protein Synthesis in Bovine Mammary Epithelial Cells

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Collage of Animal Science, Inner Mongolia Agriculture University, Hohhot 010018, China

Received date: 2018-01-25

Online published: 2018-08-18

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Abstract

The objective of this study was to determine the effects of lysine (Lys) on gene expressions and protein phosphorylation involved in milk protein synthesis in bovine mammary epithelial cells (BMECs). The 3rd generation BMECs were randomly divided into six group with six replicates per group, cells in different groups were cultured in medium with 0.5 (control), 1.0, 2.0, 4.0, 8.0 and 16.0 mmol/L Lys, respectively. After 48 h cultivation at 37℃ and 5% CO₂, adenosine triphosphate (ATP) content was measured by the method of chemiluminescence, expression levels of genes involved in milk protein synthesis were determined by real-time PCR, and phosphorylation levels of proteins involved in milk protein synthesis were determined by Western blotting method. The results showed as follows:with the increase of Lys concentration, ATP content tended to be quadratically significantly increased (P=0.050); expression levels of κ-casein (CSN3) (P=0.093), mammalian target of rapamycin (mTOR) (P=0.005), eukaryotic initiation factor 4E (eIF4E) (P=0.076) and adenosine 5'-mono-phpsphate-active protein kinase α1 (AMPKα1) genes (P=0.045) were significantly or tended to significantly quadratically changed, all showed firstly increased then decreased; expression level of α_s1-casein (CSN1S1) gene (P=0.081), phosphorylation levels of mTOR (P=0.038) and ribosomal protein S6 kinase 1 (S6K1) (P=0.022) were significantly or tend to significantly linearly decreased; phosphorylation level of adenosine 5'-mono-phpsphate-active protein kinase (AMPK) was significantly linearly increased (P=0.014). The results of analysis of variance showed that the supplementation of Lys had significantly effects on ATP content, expression levels of genes involved in milk protein synthesis and phosphorylation level of eIF4E (P<0.05),among them, the improvement effects of 2.0 to 16.0 mmol/L Lys for ATP content, 1.0 to 2.0 mmol/L Lys for CSN1S1, β-casein (CSN2), signal transducer and activator of transcription 5 (STAT5)gene expression levels, 1.0 to 8.0 mmol/L Lys for mTOR gene expression level, 1.0 to 4.0 mmol/L Lys for CSN3 gene expression level, 1.0 to 16.0 mmol/L Lys for Janus kinase 2 (JAK2) gene expression level, 2.0 mmol/L Lys for S6K1 gene expression level, 2.0 to 8.0 mmol/L Lys for eIF4E gene expression level, 2.0 to 4.0 mmol/L Lys for eIF4E phosphorylation level were better, however, 16.0 mmol/L Lys had inhibition effects on expression levels of CSN1S1, CSN3, STAT5 and mTOR genes, and 1.0 to 16.0 mmol/L Lys had inhibition effect on expression level of eukaryotic initiation 4E binding protein 1 (4EBP1) gene. In conclusion, the optimal Lys concentration for gene expressions involved in milk protein synthesis in BMECs is 1.0 to 2.0 mmol/L.

Key words： dairy cow; bovine mammary epithelial cell; lysine; milk protein

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CHEN Lu, ZHAO Yanli, GUO Xiaoyu, SHI Binlin, YAN Sumei . Effects of Lysine on Gene Expressions and Protein Phosphorylation Involved in Milk Protein Synthesis in Bovine Mammary Epithelial Cells[J]. Chinese Journal of Animal Nutrition, 2018 , 30(8) : 3142 -3150 . DOI: 10.3969/j.issn.1006-267x.2018.08.032

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