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Amino acids regulate mTOR pathway and milk protein synthesis in a mouse mammary epithelial cell line is partly mediated by T1R1/T1R3

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Abstract

Purpose

The mechanism of dietary amino acids in regulating milk protein synthesis at the translational level is not well understood. Numerous studies have shown that the amino acid signal is transferred through the mammalian target of rapamycin (mTOR) pathway; however, the extracellular amino acid-sensing mechanism that activates mTOR complex 1 is unknown. We tested the hypotheses that the T1R1/T1R3 heterodimer functions as a direct sensor of the fed state and amino acid availability preceding the mTOR pathway and affects milk protein synthesis in mammary epithelial cells.

Methods

The expression of T1R1 was repressed by T1R1 siRNA in mouse mammary epithelial cells model (HC11). Western blot was used to analyze activity of the mTOR pathway and β-casein expression, and quantitative real-time RT-PCR was used to analyze the change in mRNA abundance of amino acid transporters.

Results

The transcripts and proteins of T1R1 and T1R3 were detected in HC11 cells and mouse mammary gland tissue. siRNA silencing of T1R1 repressed β-casein synthesis in HC11 cells both with and without essential amino acids present in the culture medium. The phosphorylation of mTOR, S6K, and 4EBP1 in T1R1 knockdown HC11 cells declined to 25, 50, and 30 %, indicating T1R1 knockdown repressed the activity of the mTOR pathway. T1R1 knockdown increased the mRNAs coding three important amino acid transporters (SLC1A5 and SLC3A2/SLC7A5). Activation of the mTOR pathway was partially repressed by T1R1 siRNA or SLC7A5/SLC3A2 inhibitor (BCH, 10 mM), and the combination of these two treatments further repressed the activity of this pathway.

Conclusion

T1R1/T1R3 serves as sensor of extracellular amino acids in mouse mammary epithelial cells and involved in milk protein synthesis regulation.

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Acknowledgments

This work was partly funded by National Natural Science Foundation of China (31472077, 31101703), Natural Science Foundation of Jiangsu Province (BK20160217), Natural Science Foundation of Xuzhou Normal University (No.14XLA06), Jiangsu Overseas Research and Training Program for University Prominent Young and Middle-aged Teachers and Presidents Project, and a Project Funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions.

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Authors and Affiliations

  1. Institute of Cellular and Molecular Biology, School of Life Science, Jiangsu Normal University, Xuzhou, 221116, Jiangsu, China

    YanHong Wang, JunQiang Liu, Hui Wu, XingTang Fang, Hong Chen & ChunLei Zhang

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  1. YanHong Wang
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  2. JunQiang Liu
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  3. Hui Wu
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Correspondence to Hong Chen or ChunLei Zhang.

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Wang, Y., Liu, J., Wu, H. et al. Amino acids regulate mTOR pathway and milk protein synthesis in a mouse mammary epithelial cell line is partly mediated by T1R1/T1R3. Eur J Nutr 56, 2467–2474 (2017). https://doi.org/10.1007/s00394-016-1282-1

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  • DOI: https://doi.org/10.1007/s00394-016-1282-1

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