Arabidopsis Lectin Receptor Kinase P2K2 Is a Second Plant Receptor for Extracellular ATP and Contributes to Innate Immunity.

In animals, extracellular ATP is a well-studied signaling molecule that is recognized by plasma membrane-localized P2-type purinergic receptors. However, in contrast, much less is known about purinergic signaling in plants. P2 receptors play critical roles in a variety of animal biological processes, including immune system regulation. The first plant purinergic receptor, Arabidopsis (Arabidopsis thaliana) P2K1 (L-type lectin receptor kinase-I.9), was shown to contribute to plant defense against bacterial, oomycete, and fungal pathogens. Here, we demonstrate the isolation of a second purinergic receptor, P2K2, by complementation of an Arabidopsis p2k1 mutant. P2K2 (LecRK-I.5) has 74% amino acid similarity to P2K1. The P2K2 extracellular lectin domain binds to ATP with higher affinity than P2K1 (dissociation constant [K d] = 44.47 ± 15.73 nm). Interestingly, p2k2 and p2k1 p2k2 mutant plants showed increased susceptibility to the pathogen Pseudomonas syringae, with the double mutant showing a stronger phenotype. In vitro and in planta studies demonstrate that P2K2 and P2K1 interact and cross-phosphorylate upon extracellular ATP treatment. Thus, similar to animals, plants possess multiple purinergic receptors.

Computational prediction and in vitro analysis of the potential ligand binding site within the extracellular ATP receptor, P2K2.

The plant extracellular ATP (eATP) receptor, P2K2, binds eATP with strong ligand affinity through its extracellular lectin domain. Ligand binding activates the intracellular kinase domain of P2K2 resulting in a variety of intracellular responses and, ultimately, increased plant immunity to invading fungal and bacterial pathogens. Here, using a computational prediction approach, we developed a tertiary structure model of the P2K2 extracellular lectin domain. In silico target docking of ATP to the P2K2-binding site predicted interaction with several residues through hydrophobic interactions and hydrogen bonding. Our confirmation of the modeling was obtained by showing that H99, R144, and S256 are key residues essential for in vitro binding of ATP by P2K2.