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Cell Commun Signal. 2019 Jun 17;17(1):66. doi: 10.1186/s12964-019-0381-z.

Novel interconnections of HOG signaling revealed by combined use of two proteomic software packages.

Author information

1
Department of Biochemistry, Max F. Perutz Laboratories, Vienna BioCenter, Vienna, Austria.
2
Children's Cancer Research Institute, St. Anna Kinderspital, Vienna, Austria.
3
Structural and Computational Biology Unit, European Molecular Biology Laboratory, Meyerhofstrasse 1, 69117, Heidelberg, Germany.
4
Current Address: Department of Molecular Sociology, Max Planck Institute of Biophysics, 60438, Frankfurt am Main, Germany.
5
Mass Spectrometry Facility, Max F. Perutz Laboratories, University of Vienna, Vienna BioCenter, Vienna, Austria.
6
Mass Spectrometry Facility, Max F. Perutz Laboratories, University of Vienna, Vienna BioCenter, Vienna, Austria. wolfgang.l.reiter@univie.ac.at.

Abstract

Modern quantitative mass spectrometry (MS)-based proteomics enables researchers to unravel signaling networks by monitoring proteome-wide cellular responses to different stimuli. MS-based analysis of signaling systems usually requires an integration of multiple quantitative MS experiments, which remains challenging, given that the overlap between these datasets is not necessarily comprehensive. In a previous study we analyzed the impact of the yeast mitogen-activated protein kinase (MAPK) Hog1 on the hyperosmotic stress-affected phosphorylome. Using a combination of a series of hyperosmotic stress and kinase inhibition experiments, we identified a broad range of direct and indirect substrates of the MAPK. Here we re-evaluate this extensive MS dataset and demonstrate that a combined analysis based on two software packages, MaxQuant and Proteome Discoverer, increases the coverage of Hog1-target proteins by 30%. Using protein-protein proximity assays we show that the majority of new targets gained by this analysis are indeed Hog1-interactors. Additionally, kinetic profiles indicate differential trends of Hog1-dependent versus Hog1-independent phosphorylation sites. Our findings highlight a previously unrecognized interconnection between Hog1 signaling and the RAM signaling network, as well as sphingolipid homeostasis.

KEYWORDS:

High-osmolarity glycerol (HOG); Hog1; Hyperosmotic stress response; Kic1; MaxQuant; Mitogen-activated protein kinase (MAPK); ORMDL; Orm2; Proteome discoverer; Proteomics; p38

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