ATF4-Mediated Upregulation of REDD1 and Sestrin2 Suppresses mTORC1 Activity during Prolonged Leucine Deprivation
ATF4-Mediated Upregulation of REDD1 and Sestrin2 Suppresses mTORC1 Activity during Prolonged Leucine Deprivation
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DOI：
10.1093/jn/nxz309
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发表时间：
2020-05-01
影响因子：
4.2
通讯作者：
Kimball, Scot R.
中科院分区：
文献类型：
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作者：
Xu, Dandan;Dai, Weiwei;Kimball, Scot R.
背景：复合物 1 (mTORC1) 中雷帕霉素 (mTOR) 的蛋白激酶靶点被氨基酸激活，进而上调合成代谢过程。在营养缺乏的条件下，例如氨基酸不足，mTORC1 活性受到抑制，自噬被激活。自噬产生的溶酶体内氨基酸重新激活 mTORC1。然而，在营养不足期间持续激活 mTORC1 可能不利于细胞稳态。因此，必须存在机制来防止自噬释放的氨基酸重新激活激酶。目的：本研究的目的是测试 mTORC1 活性是否在长期亮氨酸剥夺期间通过 ATF4 依赖性上调在发育和 DNA 损伤反应 1 (REDD1) 和 Sestrin2 中调节的 mTORC1 抑制因子而受到抑制。方法：将小鼠（8 周龄；C57Bl/6 x 129SvEV）作为食物剥夺（FD）过夜，第二天早上补充一半。缺乏 ATF4、REDD1 和/或 Sestrin2 的小鼠胚胎成纤维细胞 (MEF) 被剥夺亮氨酸 016 小时。评估肝脏和细胞裂解物中的 mTORC1 活性以及 ATF4、REDD1 和 Sestrin2 表达。结果：重新喂养 FD 小鼠导致 mTORC1 激活，并与肝脏中 REDD1 和 Sestrin2 表达抑制相关。在培养的细胞中，mTORC1 对亮氨酸剥夺表现出三相反应，首先受到抑制，然后在 2 至 4 小时内短暂重新激活，并在 8 小时后再次受到抑制。再抑制与 ATF4、REDD1 和 Sestrin2 表达上调同时发生。然而，在缺乏 ATF4 的细胞中，REDD1 和 Sestrin2 的表达均不会因亮氨酸剥夺而上调，并且不存在 mTORC1 的重新抑制。此外，在缺乏 REDD1 或 Sestrin2 的细胞中，mTORC1 的再抑制减弱，而在缺乏这两种蛋白的细胞中，再抑制进一步减弱。结论：结果表明，亮氨酸剥夺以 ATF4 依赖性方式上调 REDD1 和 Sestrin2 的表达，并且这两种蛋白的表达上调参与了长期亮氨酸剥夺期间 mTORC1 的再抑制。
Background: The protein kinase target of rapamycin (mTOR) in complex 1 (mTORC1) is activated by amino acids and in turn upregulates anabolic processes. Under nutrient-deficient conditions, e.g., amino acid insufficiency, mTORC1 activity is suppressed and autophagy is activated. Intralysosomal amino acids generated by autophagy reactivate mTORC1. However, sustained mTORC1 activation during periods of nutrient insufficiency would likely be detrimental to cellular homeostasis. Thus, mechanisms must exist to prevent amino acids released by autophagy from reactivating the kinase.Objective: The objective of the present study was to test whether mTORC1 activity is inhibited during prolonged leucine deprivation through ATF4-dependent upregulation of the mTORC1 suppressors regulated in development and DNA damage response 1 (REDD1) and Sestrin2.Methods: Mice (8 wk old; C57Bl/6 x 129SvEV) were food deprived (FD) overnight and one-half were refed the next morning. Mouse embryo fibroblasts (MEFs) deficient in ATF4, REDD1, and/or Sestrin2 were deprived of leucine for 016 h. mTORC1 activity and ATF4, REDD1, and Sestrin2 expression were assessed in liver and cell lysates.Results: Refeeding FD mice resulted in activation of mTORC1 in association with suppressed expression of both REDD1 and Sestrin2 in the liver. In cells in culture, mTORC1 exhibited a triphasic response to leucine deprivation, with an initial suppression followed by a transient reactivation from 2 to 4 h and a subsequent resuppression after 8 h. Resuppression occurred concomitantly with upregulated expression of ATF4, REDD1, and Sestrin2. However, in cells lacking ATF4, neither REDD1 nor Sestrin2 expression was upregulated by leucine deprivation, and resuppression of mTORC1 was absent. Moreover, in cells lacking either REDD1 or Sestrin2, mTORC1 resuppression was attenuated, and in cells lacking both proteins resuppression was further blunted.Conclusions: The results suggest that leucine deprivation upregulates expression of both REDD1 and Sestrin2 in an ATF4-dependent manner, and that upregulated expression of both proteins is involved in resuppression of mTORC1 during prolonged leucine deprivation.