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J Proteome Res. 2015 Jul 2;14(7):2906-14. doi: 10.1021/acs.jproteome.5b00236. Epub 2015 Jun 2.

Signal Transduction Reaction Monitoring Deciphers Site-Specific PI3K-mTOR/MAPK Pathway Dynamics in Oncogene-Induced Senescence.

Author information

1
†Biomolecular Mass Spectrometry and Proteomics, Utrecht Institute for Pharmaceutical Sciences and Bijvoet Center for Biomolecular Research, Utrecht University, Padualaan 8, 3584 CH Utrecht, The Netherlands.
2
‡Netherlands Proteomics Centre, Padualaan 8, 3584 CH Utrecht, The Netherlands.
3
§Division of Molecular Oncology, The Netherlands Cancer Institute, Plesmanlaan 121, 1066 CX Amsterdam, The Netherlands.

Abstract

We report a straightforward strategy to comprehensively monitor signal transduction pathway dynamics in mammalian systems. Combining targeted quantitative proteomics with highly selective phosphopeptide enrichment, we monitor, with great sensitivity, phosphorylation dynamics of the PI3K-mTOR and MAPK signaling networks. Our approach consists of a single enrichment step followed by a single targeted proteomics experiment, circumventing the need for labeling and immune purification while enabling analysis of selected phosphorylation nodes throughout signaling pathways. The need for such a comprehensive pathway analysis is illustrated by highlighting previously uncharacterized phosphorylation changes in oncogene-induced senescence, associated with diverse biological phenotypes and pharmacological intervention of the PI3K-mTOR pathway.

KEYWORDS:

ERK1/2; MAPK; PI3K; RPS6; SRM; Ti4+-IMAC; mTOR; network biology; pathway; phosphoproteomics; signal transduction

PMID:
26011226
DOI:
10.1021/acs.jproteome.5b00236
[Indexed for MEDLINE]

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