MTORC1 promotes T-bet phosphorylation to regulate Th1 differentiation

Olesya Chornoguz, Robert S. Hagan, Azeb Haile, Matthew L. Arwood, Christopher J. Gamper, Arnob Banerjee, Jonathan D. Powell

Research output: Research - peer-reviewArticle

Abstract

CD4+ T cells lacking the mTORC1 activator Rheb fail to secrete IFN-g under Th1 polarizing conditions. We hypothesized that this phenotype is due to defects in regulation of the canonical Th1 transcription factor T-bet at the level of protein phosphorylation downstream of mTORC1. To test this hypothesis, we employed targeted mass-spectrometry proteomic analysis-multiple reaction monitoring mass spectrometry. We used this method to detect and quantify predicted phosphopeptides derived from T-bet. By analyzing activated murine wild-type and Rheb-deficient CD4+ T cells, as well as murine CD4+ T cells activated in the presence of rapamycin, a pharmacologic inhibitor of mTORC1, we were able to identify six T-bet phosphorylation sites. Five of these are novel, and four sites are consistently dephosphorylated in both Rheb-deficient CD4+ T cells and T cells treated with rapamycin, suggesting mTORC1 signaling controls their phosphorylation. Alanine mutagenesis of each of the six phosphorylation sites was tested for the ability to impair IFN-g expression. Single phosphorylation site mutants still support induction of IFN-g expression; however, simultaneous mutation of three of the mTORC1-dependent sites results in significantly reduced IFN-g expression. The reduced activity of the triple mutant T-bet is associated with its failure to recruit chromatin remodeling complexes to the Ifng gene promoter. These results establish a novel mechanism by which mTORC1 regulates Th1 differentiation, through control of T-bet phosphorylation.

LanguageEnglish (US)
Pages3939-3948
Number of pages10
JournalJournal of Immunology
Volume198
Issue number10
DOIs
StatePublished - May 15 2017

Fingerprint

Phosphorylation
mechanistic target of rapamycin complex 1
T-Lymphocytes
Sirolimus
Mass Spectrometry
Phosphopeptides
Chromatin Assembly and Disassembly
Mutagenesis
Alanine
Proteomics
Phenotype
Mutation
Genes
Proteins
T-box transcription factor TBX21

ASJC Scopus subject areas

  • Immunology

Cite this

Chornoguz, O., Hagan, R. S., Haile, A., Arwood, M. L., Gamper, C. J., Banerjee, A., & Powell, J. D. (2017). MTORC1 promotes T-bet phosphorylation to regulate Th1 differentiation. Journal of Immunology, 198(10), 3939-3948. DOI: 10.4049/jimmunol.1601078

MTORC1 promotes T-bet phosphorylation to regulate Th1 differentiation. / Chornoguz, Olesya; Hagan, Robert S.; Haile, Azeb; Arwood, Matthew L.; Gamper, Christopher J.; Banerjee, Arnob; Powell, Jonathan D.

In: Journal of Immunology, Vol. 198, No. 10, 15.05.2017, p. 3939-3948.

Research output: Research - peer-reviewArticle

Chornoguz, O, Hagan, RS, Haile, A, Arwood, ML, Gamper, CJ, Banerjee, A & Powell, JD 2017, 'MTORC1 promotes T-bet phosphorylation to regulate Th1 differentiation' Journal of Immunology, vol 198, no. 10, pp. 3939-3948. DOI: 10.4049/jimmunol.1601078
Chornoguz O, Hagan RS, Haile A, Arwood ML, Gamper CJ, Banerjee A et al. MTORC1 promotes T-bet phosphorylation to regulate Th1 differentiation. Journal of Immunology. 2017 May 15;198(10):3939-3948. Available from, DOI: 10.4049/jimmunol.1601078
Chornoguz, Olesya ; Hagan, Robert S. ; Haile, Azeb ; Arwood, Matthew L. ; Gamper, Christopher J. ; Banerjee, Arnob ; Powell, Jonathan D./ MTORC1 promotes T-bet phosphorylation to regulate Th1 differentiation. In: Journal of Immunology. 2017 ; Vol. 198, No. 10. pp. 3939-3948
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