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mTORC1 Promotes T-bet Phosphorylation To Regulate Th1 Differentiation.

J Immunol.. 2017-05; 
Chornoguz O,,,Hagan RS,,,Haile A,,Arwood ML,,Gamper CJ,,Banerjee A,Powell JD,.
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Mutagenesis Services ... Mutagenesis was verified by sequencing. Triple mutant creation and sequencing verification was con- ducted by GenScript. EL4 transfection EL4 cells were maintained in RPMI with 10% FBS and antibiotics, subsequently noted as EL4 media. … Get A Quote

摘要

CD4+ T cells lacking the mTORC1 activator Rheb fail to secrete IFN-γ 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 pho... More

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