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Two cysteine-rich receptor-like protein kinases, CRK7 and CRK43, are required for CERK1-4 dependent cell death responses in Arabidopsis thaliana

dc.contributor.advisorPetutschnig, Elena K Dr.
dc.contributor.authorTrippel, Christine
dc.titleTwo cysteine-rich receptor-like protein kinases, CRK7 and CRK43, are required for CERK1-4 dependent cell death responses in Arabidopsis thalianade
dc.contributor.refereeLipka, Volker Prof. Dr.
dc.description.abstractengPlants recognize potential pathogens via conserved molecular patterns. These are perceived by plasma membrane-associated receptor complexes leading to downstream signaling and activation of the plant immune system. In Arabidopsis thaliana, the LysM containing kinase CERK1 is an essential part of the receptor for chitin, a major component of fungal cell walls. Besides full length CERK1, a shorter N-terminal fragment is present, that is generated by ectodomain shedding and released into the apoplast. This N-terminal fragment cannot be detected in plants carrying a CERK1 mutant version with an amino acid exchange in the ectodomain (CERK1-4). Furthermore, cerk1-4 mutants show a deregulated salicylic acid dependent cell death response upon infection with powdery mildews and in senescent plants, whereas chitin responses are WT-like. The role of the shed ectodomain and the mechanism underlying the deregulated cell death response are still unclear. To identify molecular components necessary for cell death induction in cerk1-4, a suppressor screen was conducted. In this work, three independent suppressor mutations were identified. In one mutant line suppression of the cerk1-4 phenotype was probably caused by an amino acid exchange in PAD4. Since PAD4 was already known to be necessary for cerk1-4 cell death induction, this mutant was not further characterized. The other suppressor mutations affected two different cysteine-rich receptor-like protein kinases, CRK7 and CRK43. CRKs are one of the largest subgroups among Arabidopsis receptor-like kinases and contain characteristic and highly conserved cysteine-rich motifs in their ectodomains. Moreover, several CRKs were shown to play a role in plant immunity and cell death regulation. CRK7 and CRK43 were confirmed as components of the cerk1-4 pathway by independent mutants and/ or complementation approaches and localization studies were conducted. In both crk7 and crk43 mutants, immune responses upon chitin and flg22 treatment were not altered compared to the wildtype. In a point mutant isolated from the cerk1-4 suppressor screen (CRK7-4), a highly conserved cysteine residue in the ectodomain is mutated. Co-immunoprecipitation analyses revealed that CRK7 is able to interact with itself whereas CRK7-4 is not. Moreover, it was shown that CRK7 localizes predominantly to the plasma membrane, while CRK7-4 is found in the ER. Structural comparisons with related proteins and proteomics data suggest that CRK7-4 is misfolded, resulting in ER retention. The phosphorylation status was investigated in CRK7 and CRK7-4 isolated from Arabidopsis or Nicotiana benthamiana plants. Strongly phosphorylated amino acids in the activation and P+1 loop of the kinase domain were identified in CRK7, but phosphorylation of these residues was strongly reduced in CRK7-4. Nevertheless, in vitro kinase activity was shown for both CRK7 and CRK7-4. A mutant in which all phosphorylateable amino acids in the activation and P+1 loop of CRK7 were exchanged for alanine was unable to autophosphorylate or to phosphorylate an artificial substrate. These findings suggest that CRK7 might be phosphorylated and activated after homomerisation. CRK7 might function as a receptor perceiving either the CERK1 ectodomain, turnover products of the CERK1-4 ectodomain, or another yet unknown signal, and subsequently mediate cell death induction in
dc.contributor.coRefereeWiermer, Marcel PD Dr.
dc.subject.engPlant Cell Biologyde
dc.subject.engCysteine-rich Receptor-like Kinases (CRKs)de
dc.subject.engCERK1-4 dependent cell deathde
dc.subject.engPlant-pathogen interactionde
dc.affiliation.instituteBiologische Fakultät für Biologie und Psychologiede
dc.subject.gokfullBiologie (PPN619462639)de

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