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Incubation of C2C12 myotubes with the phopholipase D (PLD) inhibitor 1‐butanol ablates contraction‐induced but not leucine‐induced signaling to p70 S6 kinase (S6K1) via phosphatidic acid (PA)
Author(s) -
Atherton Philip James,
Loughna Paul,
Hill Derek Samuel,
Rankin Debbie,
Rennie Michael
Publication year - 2010
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.24.1_supplement.331.3
Subject(s) - p70 s6 kinase 1 , phosphorylation , mtorc1 , phosphatidic acid , signal transduction , biochemistry , kinase , microbiology and biotechnology , biology , leucine , chemistry , amino acid , protein kinase b , membrane , phospholipid
Signaling through mammalian target of rapamycin complex 1 (mTORC1) regulates muscle protein synthesis (MPS) responses to amino acids (AA) (particularly leucine, (Leu)) and contraction via mechanisms independent of upstream insulin signaling (e.g. phosphoinositide 3‐kinase). Recently, it was shown that PLD‐dependent production of the lipid 2 nd ‐messenger PA (which binds and relieves inhibition of mTORC1 by FK506‐binding protein 12, thus activating mTORC1) is necessary for signaling to S6K1 after eccentric contractions. We hypothesized that, similarly, Leu‐induced increases in S6K1 signaling might also be PA‐dependent. Murine C2C12 myotubes were contracted for 30 min (1 or 4 Hz, 20 V, 24 ms pulses) or incubated with 1 mM Leu (n=6) in the presence/absence of 0.25 % 1‐butanol, a potent inhibitor of PA production by PLD. Phosphorylation of S6K1 Thr389 was measured by immunoblotting. Increased phosphorylation of S6K1 with 4 Hz stimulation exceeded that with 1 Hz (+53 vs. +33%; P <0.05), but was ablated by 1‐butanol at both 1 and 4 Hz. Conversely, the increase in S6K1 phosphorylation caused by Leu (+33%; P <0.01) was unaffected by 1‐butanol (+31%; P <0.01), which also did not alter basal S6K1 phosphorylation. Thus, whereas contraction requires PA for signaling to S6K1, Leu regulates mTORC1 signaling separately, perhaps explaining why AA and contraction synergistically stimulate MPS. Support: BBSRC, RCUK, EXGENESIS Grant Funding Source: Physiology (APS)