DGK5ß-derived phosphatidic acid regulates ROS production in plant immunity by stabilizing NADPH oxidase.

In plant immunity, phosphatidic acid (PA) regulates reactive oxygen species (ROS) by binding to respiratory burst oxidase homolog D (RBOHD), an NADPH oxidase responsible for ROS production. Here, we analyze the influence of PA binding on RBOHD activity and the mechanism of RBOHD-bound PA generation. PA binding enhances RBOHD protein stability by inhibiting vacuolar degradation, thereby increasing chitin-induced ROS production. Mutations in diacylglycerol kinase 5 (DGK5), which phosphorylates diacylglycerol to produce PA, impair chitin-induced PA and ROS production. The DGK5 transcript DGK5ß (but not DGK5α) complements reduced PA and ROS production in dgk5-1 mutants, as well as resistance to Botrytis cinerea. Phosphorylation of S506 residue in the C-terminal calmodulin-binding domain of DGK5ß contributes to the activation of DGK5ß to produce PA. These findings suggest that DGK5ß-derived PA regulates ROS production by inhibiting RBOHD protein degradation, elucidating the role of PA-ROS interplay in immune response regulation.

Reversible phosphorylation of a lectin-receptor-like kinase controls xylem immunity.

Pattern-recognition receptors (PRRs) mediate basal resistance to most phytopathogens. However, plant responses can be cell type specific, and the mechanisms governing xylem immunity remain largely unknown. We show that the lectin-receptor-like kinase LORE contributes to xylem basal resistance in Arabidopsis upon infection with Ralstonia solanacearum, a destructive plant pathogen that colonizes the xylem to cause bacterial wilt. Following R. solanacearum infection, LORE is activated by phosphorylation at residue S761, initiating a phosphorelay that activates reactive oxygen species production and cell wall lignification. To prevent prolonged activation of immune signaling, LORE recruits and phosphorylates type 2C protein phosphatase LOPP, which dephosphorylates LORE and attenuates LORE-mediated xylem immunity to maintain immune homeostasis. A LOPP knockout confers resistance against bacterial wilt disease in Arabidopsis and tomatoes without impacting plant growth. Thus, our study reveals a regulatory mechanism in xylem immunity involving the reversible phosphorylation of receptor-like kinases.

Tomato LysM receptor kinase 4 mediates chitin-elicited fungal resistance in both leaves and fruit.

Fungal infection is a major cause of crop and fruit losses. Recognition of chitin, a component of fungal cell walls, endows plants with enhanced fungal resistance. Here, we found that mutation of tomato LysM receptor kinase 4 (SlLYK4) and chitin elicitor receptor kinase 1 (SlCERK1) impaired chitin-induced immune responses in tomato leaves. Compared with the wild type, sllyk4 and slcerk1 mutant leaves were more susceptible to Botrytis cinerea (gray mold). SlLYK4 extracellular domain showed strong binding affinity to chitin, and the binding of SlLYK4 induced SlLYK4-SlCERK1 association. Remarkably, qRT-PCR analysis indicated that SlLYK4 was highly expressed in tomato fruit, and ß-GLUCURONIDASE (GUS) expression driven by the SlLYK4 promoter was observed in tomato fruit. Furthermore, SlLYK4 overexpression enhanced disease resistance not only in leaves but also in fruit. Our study suggests that chitin-mediated immunity plays a role in fruit, providing a possible way to reduce fungal infection-related fruit losses by enhancing the chitin-induced immune responses.

Tomato receptor-like cytosolic kinase RIPK confers broad-spectrum disease resistance without yield penalties.

Production of reactive oxygen species (ROS) is an important immune response in plant multilayer defense mechanisms; however, direct modification of ROS homeostasis to breed plants with broad-spectrum resistance to disease has not yet been successful. In Arabidopsis, the receptor-like cytosolic kinase AtRIPK regulates broad-spectrum ROS signaling in multiple layers of the plant immune system. Upon treatment with immune elicitors, AtRIPK is activated and phosphorylates nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, which leads to ROS production. In this study, we identified an AtRIPK ortholog in tomatoes and generated knockdown mutants using CRISPR/Cas9 technology. Slripk mutants displayed reduced ROS production in response to representative immune elicitors and were susceptible to pathogenic bacteria and fungi from different genera, including Ralstonia solanacearum, Pectobacterium carotovorum, Botrytis cinerea, and Fusarium oxysporum, which are leaf and root pathogens with hemibiotrophic and necrotrophic infection strategies. In contrast, transgenic tomato plants overexpressing SlRIPK are more resistant to these pathogens. Remarkably, the slripk mutants and SlRIPK-overexpressing transgenic plants did not exhibit significant growth retardation or yield loss. These results suggest that overexpression of SlRIPK confers broad-spectrum disease resistance without a yield penalty in tomato plants. Our findings suggest that modifying ROS homeostasis by altering the regulatory components of ROS production in plant immunity could contribute to engineering or breeding broad-spectrum disease-resistant crops without yield penalty.