Fuels for ROS signaling in plant immunity

Mitogen-activated protein kinase phosphatase 1 controls broad spectrum disease resistance in Arabidopsis thaliana through diverse mechanisms of immune activation

“…MKP1 was shown to function as a negative regulator of MAMP-dependent ROS production, since mkp1 mutant alleles ( mkp1-1 and mkp1-2 ) displayed enhanced ROS accumulation in response to flg22 and chitin ( Escudero et al., 2019 ). Considering that induction of RBOHD, the key oxidase responsible of these ROS, is mainly achieved by its phosphorylation by different kinases ( Castro et al., 2021 ; Wu et al., 2023 ), we hypothesized that MKP1 could antagonize with some of these regulatory kinases and dephosphorylate RBOHD to limit ROS production. To assess this hypothesis, we introduced in mkp1-1 rbohD and rbohD genetic backgrounds constructs harboring RBOHD wild-type (WT) gene fused to a tag (FLAG) under the control of its own promoter ( pD::FLAG-RBOHD line), or RBHOD carrying mutations in the phosphosites of RBOHD that are the targets of activating kinases during immunity: Ser (S) to Ala (A) mutations in three phosphosites that are the target of BIK1 during PTI responses ( pD::FLAG::RBOHD S39A/S339A/S343A ; Kadota et al., 2014 ; Li et al., 2014 ) and in two phosphosites that are also activated in PTI and ETI responses ( pD::FLAG::RBOHD S343A/S347A ; Kadota et al., 2019 ).…”

“…We also performed the same experiments after elicitation of all these genotypes with the fungal MAMP chitohexaose (CHI6), obtaining similar results, though the effect of mkp1-1 mutation on ROS levels was lower in pD::FLAG::RBOHD S343A/S347A than in pD::FLAG::RBOHD S39A/S339A/S343A ( Figure 1B and Supplementary Figure S1B ). The fact that the RBOHD lines with altered phosphosites displayed enhanced ROS production under mkp1-1 background compared to WT ( MKP1 ) plants suggests that MKP1 could negatively regulate RBOHD by mediating dephosphorylation of some additional phosphosites to the ones that are targeted by BIK1 and the main regulatory kinases of RBOHD documented ( Castro et al., 2021 ; Wu et al., 2023 ).…”

“…One critical aspect of the regulation of defense responses is the control of ROS production. ROS are produced during early PTI responses and are potentiated through ETI to a second wave with more sustained ROS accumulation ( Castro et al., 2021 ; Wu et al., 2023 ). NADPH oxidases, called in plants RESPIRATORY BUST OXIDASE HOMOLOGS (RBOHs), are the main enzymes that account for most of these pathogen dependent ROS production ( Torres and Dangl, 2005 ).…”

“…The activation of this oxidase has been well characterized and involves multiple posttranslational modifications mainly acting at the cytosolic N-terminal domain of the protein, which contains Ca 2+ binding EF hands ( Castro et al., 2021 ). Upon pathogen recognition, several kinases act in concert at this N-terminal domain to regulate RBOHD activation ( Kadota et al., 2015 ; Castro et al., 2021 ; Wu et al., 2023 ). These kinases that phosphorylate RBOHD N-terminal include: i) receptor-like cytoplasmic kinases (RLCKs) like BOTRYTIS INDUCED KINASE 1 (BIK1; Kadota et al., 2014 ), which plays a preeminent role in RBOHD regulation, and the RESISTANCE TO PSEUDOMONAS SYRINGAE PV.…”

“…The precise orchestration of RBOHD activation involves convergent phosphorylation events at some specific RBOHD Ser/Tyr residues (e.g. Ser343 and Ser347) by these kinases, contributing to the fine-tuning of ROS production in response to different stimuli ( Wu et al., 2023 ). Furthermore, phosphorylation of C-terminal residues by receptors like CYSTEINE-RICH RECEPTOR KINASE 2 (CRK2) and persulfidation of specific Cys in the C-terminus also contribute to the activation of AtRBOHD-dependent ROS production, emphasizing the complexity and versatility of regulatory mechanisms governing plant immunity mediated by RBOHD ( Kimura et al., 2020 ; Shen et al., 2020 ).…”