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1. Jasmonate Primes Plant Responses to Extracellular ATP

   https://doi.org/10.1101/2024.11.07.622526  

   Jewell, Jeremy B; Carlton, Ashleigh S; Tolley, Jordan P; Bartley, Laura E; Tanaka, Kiwamu (September 2025, The Plant Journal)

   Extracellular ATP (eATP) signaling inArabidopsis thalianais mediated by the purinoceptor P2K1. Previous studies have clarified that the downstream transcriptional responses to eATP involve jasmonate (JA)-based signaling components such as the JA receptor (COI1) and JA-responsive bHLH transcription factors (MYCs). However, the specific contributions of JA signaling itself on eATP signaling are unexplored. Here, we report that JA primes plant responses to eATP through P2K1. Our findings show that JA treatment significantly upregulatesP2K1transcription, corroborating our observation that JA facilitates eATP-induced cytosolic calcium elevation and transcriptional reprogramming in a JA signaling-dependent manner. Additionally, we find that salicylic acid pretreatment represses eATP-induced plant response. These results suggest that JA accumulation during biotic or abiotic stresses may potentiate eATP signaling, enabling plants to better cope with subsequent stress events. Plant hormone jasmonate (JA) enhances plant responses to extracellular ATP (eATP) inArabidopsis thalianathrough a mechanism dependent on the JA receptor COI1 and the eATP receptor P2K1. The reciprocal amplification of these signals provides a mechanistic explanation for how plants effectively respond to different stress events. 

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2. Cyclic nucleotide‐gated ion channel 6 is involved in extracellular ATP signaling and plant immunity

   https://doi.org/10.1111/tpj.15636  

   Duong, Ha N.; Cho, Sung‐Hwan; Wang, Limin; Pham, An Q.; Davies, Julia M.; Stacey, Gary (December 2021, The Plant Journal)

   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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3. The Raf-like MAPKKK INTEGRIN-LINKED KINASE 5 regulates purinergic receptor-mediated innate immunity in Arabidopsis

   https://doi.org/10.1093/plcell/koad029  

   Kim, Daewon; Chen, Dongqin; Ahsan, Nagib; Jorge, Gabriel Lemes; Thelen, Jay J.; Stacey, Gary (February 2023, The Plant Cell)

   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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4. Activation of the plant mevalonate pathway by extracellular ATP

   https://doi.org/10.1038/s41467-022-28150-w  

   Cho, Sung-Hwan; Tóth, Katalin; Kim, Daewon; Vo, Phuc H.; Lin, Chung-Ho; Handakumbura, Pubudu P.; Ubach, Albert Rivas; Evans, Sterling; Paša-Tolić, Ljiljana; Stacey, Gary (December 2022, Nature Communications)

   Abstract The mevalonate pathway plays a critical role in multiple cellular processes in both animals and plants. In plants, the products of this pathway impact growth and development, as well as the response to environmental stress. A forward genetic screen of Arabidopsis thaliana using Ca 2+ -imaging identified mevalonate kinase (MVK) as a critical component of plant purinergic signaling. MVK interacts directly with the plant extracellular ATP (eATP) receptor P2K1 and is phosphorylated by P2K1 in response to eATP. Mutation of P2K1-mediated phosphorylation sites in MVK eliminates the ATP-induced cytoplasmic calcium response, MVK enzymatic activity, and suppresses pathogen defense. The data demonstrate that the plasma membrane associated P2K1 directly impacts plant cellular metabolism by phosphorylation of MVK, a key enzyme in the mevalonate pathway. The results underline the importance of purinergic signaling in plants and the ability of eATP to influence the activity of a key metabolite pathway with global effects on plant metabolism. 

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5. ORA47 is a transcriptional regulator of a general stress response hub

   https://doi.org/10.1111/tpj.15688  

   Zeng, Liping; Chen, Hao; Wang, Yaqi; Hicks, Derrick; Ke, Haiyan; Pruneda‐Paz, Jose; Dehesh, Katayoon (February 2022, The Plant Journal)

   SUMMARY Transcriptional regulators of the general stress response (GSR) reprogram the expression of selected genes to transduce informational signals into cellular events, ultimately manifested in a plant's ability to cope with environmental challenges. Identification of the core GSR regulatory proteins will uncover the principal modules and their mode of action in the establishment of adaptive responses. To define the GSR regulatory components, we employed a yeast‐one‐hybrid assay to identify the protein(s) binding to the previously established functional GSR motif, termed the rapid stress response element (RSRE). This led to the isolation of octadecanoid‐responsive AP2/ERF‐domain transcription factor 47 (ORA47), a methyl jasmonate inducible protein. Subsequently, ORA47 transcriptional activity was confirmed using the RSRE‐driven luciferase (LUC) activity assay performed in the ORA47 loss‐ and gain‐of‐function lines introgressed into the 4xRSRE::Luc background. In addition, the prime contribution of CALMODULIN‐BINDING TRANSCRIPTIONAL ACTIVATOR3 (CAMTA3) protein in the induction of RSRE was reaffirmed by genetic studies. Moreover, exogenous application of methyl jasmonate led to enhanced levels ofORA47andCAMTA3transcripts, as well as the induction of RSRE::LUC activity. Metabolic analyses illustrated the reciprocal functional inputs of ORA47 and CAMTA3 in increasing JA levels. Lastly, transient assays identified JASMONATE ZIM‐domain1 (JAZ1) as a repressor of RSRE::LUC activity. Collectively, the present study provides fresh insight into the initial features of the mechanism that transduces informational signals into adaptive responses. This mechanism involves the functional interplay between the JA biosynthesis/signaling cascade and the transcriptional reprogramming that potentiates GSR. Furthermore, these findings offer a window into the role of intraorganellar communication in the establishment of adaptive responses. 

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