Myogenic differentiation is dependent on both the kinase function and the N-terminal sequence of mammalian target of rapamycin

Lili Shu, Xiongwen Zhang, Peter J. Houghton

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Abstract

The mammalian target of rapamycin (mTOR) is a serine/threonine protein kinase known to control initiation of translation through two downstream pathways: eukaryotic initiation factor 4E-binding protein 1 (4E-BP1)/eukaryotic initiation factor 4E and ribosomal p70 S6 kinase (S6K1). We previously showed in C2C12 murine myoblasts that rapamycin arrests cells in G1 phase and completely inhibits terminal myogenesis. To elucidate the pathways that regulate myogenesis, we established stable C2C12 cell lines that express rapamycin-resistant mTOR mutants (mTORrr; S2035I) that have N-terminal deletions (Δ10 or Δ91) or are full-length kinase-dead mTORrr proteins. Additional clones expressing a constitutively active S6K1 were also studied. Our results show that Δ10mTORrr signals 4E-BP1 and permits rapamycin-treated myoblasts to differentiate, confirming the mTOR dependence of the inhibition of myogenesis by rapamycin. C2C12 cells expressing either Δ91mTORrr or kinase-dead mTORrr(D2338A) could not phosphorylate 4E-BP1 in the presence of rapamycin and could not abrogate the inhibition of myogenesis. Taken together, our results indicate that both the kinase function of mTOR and the N terminus (residues 11-91, containing part of the first HEAT domain) are essential for myogenic differentiation. In contrast, constitutive activation of S6K1 does not abrogate rapamycin inhibition of either proliferation or myogenic differentiation.

Original languageEnglish (US)
Pages (from-to)16726-16732
Number of pages7
JournalJournal of Biological Chemistry
Volume277
Issue number19
DOIs
StatePublished - May 10 2002

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ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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