Amino acids regulate mTOR pathway and milk protein synthesis in a mouse mammary epithelial cell line is partly mediated by T1R1/T1R3
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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小鼠乳腺上皮细胞系中氨基酸调节 mTOR 通路和乳蛋白合成部分由 T1R1/T1R3 介导
DOI：
10.1007/s00394-016-1282-1
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发表时间：
2017-12
期刊：
影响因子：
--
通讯作者：
Zhang C
中科院分区：
文献类型：
--
作者：
Wang Y;Liu J;Wu H;Fang X;Chen H;Zhang C
目的饲粮氨基酸在翻译水平上调控乳蛋白合成的机制尚不清楚。大量研究表明，氨基酸信号是通过哺乳动物雷帕霉素靶蛋白（mTOR）途径传递的；然而，激活mTOR复合体1的细胞外氨基酸感应机制尚不清楚。我们验证了T1R1/T1R3异源二聚体在mTOR通路之前作为喂养状态和氨基酸可用性的直接传感器，并影响乳腺上皮细胞中乳蛋白合成的假设。方法用T1R1 siRNA抑制小鼠乳腺上皮细胞模型（HC11）中T1R1的表达。Western blot检测mTOR通路活性及β-酪蛋白表达，实时荧光定量RT-PCR检测氨基酸转运蛋白mRNA丰度变化。结果在HC11细胞和小鼠乳腺组织中均检测到T1R1和T1R3的转录本和蛋白。siRNA沉默T1R1抑制了HC11细胞中β-酪蛋白的合成，无论培养基中是否存在必需氨基酸。在T1R1敲低的HC11细胞中，mTOR、S6K和4EBP1的磷酸化分别下降到25%、50%和30%，表明T1R1敲低抑制了mTOR通路的活性。T1R1敲低增加了编码三种重要氨基酸转运体（SLC1A5和SLC3A2/SLC7A5）的mrna。mTOR通路的激活被T1R1 siRNA或SLC7A5/SLC3A2抑制剂（BCH, 10 mM）部分抑制，这两种处理的联合进一步抑制了该通路的活性。结论t1r1 /T1R3是小鼠乳腺上皮细胞胞外氨基酸的传感器，参与乳蛋白的合成调控。
PurposeThe 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.MethodsThe 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.ResultsThe 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.ConclusionT1R1/T1R3 serves as sensor of extracellular amino acids in mouse mammary epithelial cells and involved in milk protein synthesis regulation.
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影响因子：
2.1
作者：
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通讯作者：
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