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Knockdown of Deptor, an mTOR binding protein, increases muscle protein synthesis
Author(s) -
Kazi Abid Abdulaziz,
HongBrown Ly,
Lang Charles H
Publication year - 2011
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.25.1_supplement.1064.3
Subject(s) - pi3k/akt/mtor pathway , p70 s6 kinase 1 , mtorc1 , microbiology and biotechnology , rptor , mtorc2 , myocyte , gene knockdown , phosphorylation , autophagy , chemistry , biology , apoptosis , signal transduction , biochemistry
Deptor is an mTOR binding protein that affects cell metabolism. The hypothesis of the present study was that knock down (KD) of Deptor in C2C12 myocytes will increase protein synthesis via stimulating mTOR‐S6K1 signaling. Deptor KD was achieved using lentiviral particles containing shRNA targeting the mouse Deptor mRNA sequence, whereas the control cells were transfected with shRNA that does not target any known mammalian sequence. KD reduced Deptor mRNA and protein content by 90%, and increased phosphorylation of mTOR kinase substrates, 4E‐BP1 and S6K1. This activation of mTOR correlated with increased rates of protein synthesis. The responsiveness of KD and control myocytes to anabolic (IGF‐I) and catabolic (AICAR) stimuli were unaltered. Deptor KD myoblasts were both larger in diameter and exhibited an increase in mean cell volume. Deptor KD cells had an increased percentage of cells in the S phase. Immunoblotting confirmed an increase in the total p21 and phosphorylation (Ser807/Ser811) of Rb protein which is critical for the G 1 ‐S phase transition in myoblasts. Deptor KD did not alter myoblast apoptosis or autophagy as evidenced by the lack of change for cleaved caspase‐3 and LC3B, respectively. Finally, in vivo Deptor KD (~50% reduction) by electroporation in C57/BL6 mice did not alter weight of control muscles but prevented atrophy produced by 3 d of immobilization. Thus our data support the hypothesis that deptor is an important regulator of protein metabolism in myocytes. (Funding support: GM38032 to CHL)