PBL13 is a serine/threonine protein kinase that negatively regulates Arabidopsis immune responses.

Abstract

Receptor-like cytoplasmic kinases (RLCKs) are a subset of plant receptor-like kinases lacking both extracellular and transmembrane domains. Some of the 46 members in the Arabidopsis (Arabidopsis thaliana) RLCK subfamily VII have been linked to plant innate immunity; however, most remain uncharacterized. Thus, multiple subfamily VII members are expected to be involved in plant immune signaling. Here, we investigate the role of AvrPphB SUSCEPTIBLE1-LIKE13 (PBL13), a subfamily VII RLCK with unique domain architecture. Unlike other characterized RLCKs, PBL13 transfer DNA insertion lines exhibit enhanced disease resistance after inoculation with virulent Pseudomonas syringae. The pbl13-2 knockout also exhibits elevated basal-level expression of the PATHOGENESIS-RELATED GENE1 defense marker gene, enhanced reactive oxygen species (ROS) burst in response to perception of bacterial microbial patterns, and accelerated flagellin-induced activation of mitogen-activated protein kinases. Recombinant PBL13 is an active kinase, and its primary autophosphorylated sites map to a 15-amino acid repeat motif unique to PBL13. Complementation of pbl13-2 with PBL13-3xFLAG converts the enhanced resistance and elevated ROS phenotypes back to wild-type levels. In contrast, kinase-dead PBL13 K111A -3xFLAG was unable to rescue pbl13-2 disease phenotypes. Consistent with the enhanced ROS burst in the pbl13-2 knockout, PBL13 is able to associate with the nicotinamide adenine dinucleotide phosphate, reduced oxidase RESPIRATORY BURST OXIDASE HOMOLOG PROTEIN D (RBOHD) by split-luciferase complementation assay, and this association is disrupted by flagellin treatment. We conclude that the PBL13 kinase negatively regulates plant innate immunity to pathogenic bacteria and can associate with RBOHD before pathogen perception. These data are consistent with the hypothesis that PBL13 acts to prevent inappropriate activation of defense responses in the absence of pathogen challenge.

A Cysteine-Rich Protein Kinase Associates with a Membrane Immune Complex and the Cysteine Residues Are Required for Cell Death

“…The highly conserved Lys (K377) residue in CRK28's ATP-binding motif was mutated to Asn (N; 35S:CRK28 K377N -FLAG). Mutating this conserved Lys residue in Ser/Thr protein kinases has been shown to block activity Lin et al, 2015). None of the leaves infiltrated with 35S: CRK28 K377N -FLAG exhibited cell death, but all leaves infiltrated with wild-type 35S:CRK28-FLAG exhibited cell death (Fig.…”

“…In addition, the transmembrane protein RBOHD, an NADPH oxidase responsible for the production of extracellular ROS, is activated through transphosphorylation by the receptor-like cytoplasmic kinase BIK1 and calcium-dependent protein kinases (Kadota et al, 2015). Mutation of the conserved Lys residue in the kinase domain of the Ser/Thr protein kinases is known to abolish kinase activity and thereby signaling Lin et al, 2015). In vitro kinase activities have been demonstrated for CRK6, CRK7, and CRK36 (Tanaka et al, 2012;Idänheimo et al, 2014).…”

Receptor Kinases in Plant-Pathogen Interactions: More Than Pattern Recognition

“…Furthermore, RbohD contains EF-hand motifs for calcium ion binding, which likely provide further regulation. XLG2 and PBL13 may also positively or negatively regulate RbohD activity, respectively, although the underlying mechanisms are unknown (Figure 3; Liang et al, 2016;Lin et al, 2015).…”

Changes in PUB22 Ubiquitination Modes Triggered by MITOGEN-ACTIVATED PROTEIN KINASE3 Dampen the Immune Response

“…LR reactions were performed with the destination vectors pCAMBIA/des/cLuc and pCAMBIA/des/nLuc (Chen et al, 2008;Lin et al, 2015b). Protoplasts were generated as previously described (Wu et al, 2009) from pub22 pub23 pub24 or mkp3 mutant plants and transformed with 25 mg/mL of each construct.…”