Sustained mitogen-activated protein kinase activation reprograms defense metabolism and phosphoprotein profile in Arabidopsis thaliana

Mitogen-activatedproteinkinases(MAPKs)targetavarietyofproteinsubstratestoregulatecellularsignalingprocessesineukaryotes.Inplants,thenumberofidentifiedMAPKsubstratesthatcontrolplantdefenseresponsesisstilllimited.Here,wegeneratedtransgenicArabidopsisthalianaplantswithaninduciblesystemtosimulateinvivoactivationoftwostress-activatedMAPKs,MPK3,andMPK6.MetabolomeanalysisrevealedthatthisartificialMPK3/6activation(withoutanyexposuretopathogensorotherstresses)issufficienttodrivetheproductionofmajordefense-relatedmetabolites,includingvariouscamalexin,indoleglucosinolateandagmatinederivatives.Anaccompanying(phospho)proteomeanalysisledtodetectionofhundredsofpotentialphosphoproteinsdownstreamofMPK3/6activation.BesidesknownMAPKsubstrates,manycandidatesonthislistpossesstypicalMAPK-targetedphosphositesandinmanycases,thecorrespondingphosphopeptidesweredetectedbymassspectrometry.Notably,severaloftheseputativephosphoproteinshavebeenreportedtobeassociatedwiththebiosynthesisofantimicrobialdefensesubstances(e.g.,WRKYtranscriptionfactorsandproteinsencodedbythegenesfromthe“PEN”pathwayrequiredforpenetrationresistancetofilamentouspathogens).Thus,thisworkprovidesaninventoryofcandidatephosphoproteins,includingputativedirectMAPKsubstrates,forfutureanalysisofMAPK-mediateddefensecontrol.(ProteomicsdataareavailablewiththeidentifierPXD001252viaProteomeXchange,http://proteomecentral.proteomexchange.org).