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. 2019 Oct 17;9:14910. doi: 10.1038/s41598-019-51410-7

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© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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SM decreases mSIN1 levels in mTORC2 by PLD2-induced PA production in C2C12 myoblasts. (A) C2C12 myoblasts were incubated either under terrestrial gravity (1 g) or under SM for 36 h, lysed, and subjected to immunoprecipitation using antibodies against either raptor or rictor, and analyzed by western blotting. (B) Cells were treated as in (A), lysed, and subjected to quantitative RT-PCR for PLD1 and PLD2 (n = 4). (C) Cells were treated as in (A), lysed, and analyzed by western blotting. (D) PLD assay was performed using a PLD assay kit (n = 4). (E) Cells were treated with or without 300 μM C8-PA for 30 min, lysed, then subjected to immunoprecipitation with antibody against rictor, and analyzed by western blotting. Abbreviations: simulated microgravity (SM); immunoprecipitate (IP); Phosphatidic acid (PA). All data are blots representative of 3 to 5 independent experiments. Data are expressed as mean ± SD, with Mann-Whitney U test performed as indicated. *P < 0.05; versus control under 1 g.