Identification of isoform-specific dynamics in phosphorylation-dependent STAT5 dimerization by quantitative mass spectrometry and mathematical modeling.


STAT5A and STAT5B are important transcription factors that dimerize and transduce activation signals of cytokine receptors directly to the nucleus. A typical cytokine that mediates STAT5 activation is erythropoietin (Epo). Differential functions of STAT5A and STAT5B have been reported. However, the extent to which phosphorylated STAT5A and STAT5B (pSTAT5A, pSTAT5B) form homo- or heterodimers is not understood, nor is how this might influence the signal transmission to the nucleus. To study this, we designed a concept to investigate the isoform-specific dimerization behavior of pSTAT5A and pSTAT5B that comprises isoform-specific immunoprecipitation (IP), measurement of the degree of phosphorylation, and isoform ratio determination between STAT5A and STAT5B. For the main analytical method, we employed quantitative label-free and -based mass spectrometry. For the cellular model system, we used Epo receptor (EpoR)-expressing BaF3 cells (BaF3-EpoR) stimulated with Epo. Three hypotheses of dimer formation between pSTAT5A and pSTAT5B were used to explain the analytical results by a static mathematical model: formation of (i) homodimers only, (ii) heterodimers only, and (iii) random formation of homo- and heterodimers. The best agreement between experimental data and model simulations was found for the last case. Dynamics of cytoplasmic STAT5 dimerization could be explained by distinct nuclear import rates and individual nuclear retention for homo- and heterodimers of phosphorylated STAT5.


PubMed ID: 25333863

Projects: SBEpo - Systems Biology of Erythropoietin

Publication type: Journal

Journal: J Proteome Res

Citation: J Proteome Res. 2014 Dec 5;13(12):5685-94. doi: 10.1021/pr5006923. Epub 2014 Nov 7.

Date Published: 5th Dec 2014

Registered Mode: Not specified

Authors: M. E. Boehm, L. Adlung, M. Schilling, S. Roth, U. Klingmuller, W. D. Lehmann

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Created: 13th Oct 2016 at 12:10

Last updated: 3rd Aug 2020 at 18:38

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