Identifizierung von MAP-Kinase-Substraten in Arabidopsis thaliana

  • Identification of MAP kinase substrates in Arabidopsis thaliana

Thomas, Martin; Conrath, Uwe; Usadel, Björn (Thesis advisor)

Aachen (2016)
Dissertation / PhD Thesis

Dissertation, RWTH Aachen, 2016

Abstract

Mitogen-activated protein kinases (MAP-kinases) are signal transduction components that are conserved in eukaryotes. They play important roles in growth and development as well as in defense against biotic and abiotic stress. They are part of a molecular mechanism of salicylic acid-dependent priming of defense responses. MAP-kinases transduce extracellular signals into cellular responses. The key step which actually turns the signal transduction into a response is performed by the substrates of the MAP-kinases. In contrast to the animal field only few MAP-Kinase substrates are known in the model plant Arabidopsis thaliana. In this work substrates of MAP-kinases were phosphor-specifically enriched by Tandem metal oxide / hydroxide affinity chromatography (Tandem-MOAC) and identified by mass spectrometry. The phosphorylated amino acids were determined in Tandem mass spectrometry (MS/MS) experiments. Quantitative analysis enabled the identification of novel putative substrates of the stress-responsive MAP-kinases MPK3 and/or MPK6. Induction of defense priming by treatment with salicylic acid allowed for the identification of putative MAP-kinase substrates involved in the priming process. Analysis of a loss-of-function mutant facilitated recording of putative substrates that are phosphorylated specifically but not necessarily exclusively by MPK3.