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SUMMARY Extracellular ATP (eATP) is known to act as a danger signal in both plants and animals. In plants, eATP is recognized by the plasma membrane (PM)‐localized receptor P2K1 (LecRK‐I.9). Among the first measurable responses to eATP addition is a rapid rise in cytoplasmic free calcium levels ([Ca2+]cyt), which requires P2K1. However, the specific transporter/channel proteins that mediate this rise in [Ca2+]cytare unknown. Through a forward genetic screen, we identified an Arabidopsis ethylmethanesulfonate (EMS) mutant impaired in the [Ca2+]cytresponse to eATP. Positional cloning revealed that the mutation resided in thecngc6gene, which encodes cyclic nucleotide‐gated ion channel 6 (CNGC6). Mutation of theCNGC6gene led to a notable decrease in the PM inward Ca2+current in response to eATP. eATP‐induced mitogen‐activated protein kinase activation and gene expression were also significantly lower incngc6mutant plants. In addition,cngc6mutant plants were also more susceptible to the bacterial pathogenPseudomonas syringae. Taken together, our results indicate that CNGC6 plays a crucial role in mediating eATP‐induced [Ca2+]cytsignaling, as well as plant immunity.
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The Raf-like MAPKKK INTEGRIN-LINKED KINASE 5 regulates purinergic receptor-mediated innate immunity in Arabidopsis
Abstract Mitogen-activated protein (MAP) kinase signaling cascades play important roles in eukaryotic defense against various pathogens. Activation of the extracellular ATP (eATP) receptor P2K1 triggers MAP kinase 3 and 6 (MPK3/6) phosphorylation, which leads to an elevated plant defense response. However, the mechanism by which P2K1 activates the MAPK cascade is unclear. In this study, we show that in Arabidopsis thaliana, P2K1 phosphorylates the Raf-like MAP kinase kinase kinase (MAPKKK) INTEGRIN-LINKED KINASE 5 (ILK5) on serine 192 in the presence of eATP. The interaction between P2K1 and ILK5 was confirmed both in vitro and in planta and their interaction was enhanced by ATP treatment. Similar to P2K1 expression, ILK5 expression levels were highly induced by treatment with ATP, flg22, Pseudomonas syringae pv. tomato DC3000, and various abiotic stresses. ILK5 interacts with and phosphorylates the MAP kinase MKK5. Moreover, phosphorylation of MPK3/6 was significantly reduced upon ATP treatment in ilk5 mutant plants, relative to wild-type (WT). The ilk5 mutant plants showed higher susceptibility to P. syringae pathogen infection relative to WT plants. Plants expressing only the mutant ILK5S192A protein, with decreased kinase activity, did not activate the MAPK cascade upon ATP addition. These results suggest that eATP activation of P2K1 results in transphosphorylation of the Raf-like MAPKKK ILK5, which subsequently triggers the MAPK cascade, culminating in activation of MPK3/6 associated with an elevated innate immune response.
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- Award ID(s):
- 2048410
- PAR ID:
- 10398344
- Publisher / Repository:
- Oxford University Press
- Date Published:
- Journal Name:
- The Plant Cell
- ISSN:
- 1040-4651
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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