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Arabidopsis thaliana CYCLIC NUCLEOTIDE-GATED CHANNEL2 mediates extracellular ATP signal transduction in root epidermis.

ABSTRACT: Damage can be signalled by extracellular ATP (eATP) using plasma membrane (PM) receptors to effect cytosolic free calcium ion ([Ca²⁺ ]_cyt ) increase as a second messenger. The downstream PM Ca²⁺ channels remain enigmatic. Here, the Arabidopsis thaliana Ca²⁺ channel subunit CYCLIC NUCLEOTIDE-GATED CHANNEL2 (CNGC2) was identified as a critical component linking eATP receptors to downstream [Ca²⁺ ]_cyt signalling in roots. Extracellular ATP-induced changes in single epidermal cell PM voltage and conductance were measured electrophysiologically, changes in root [Ca²⁺ ]_cyt were measured with aequorin, and root transcriptional changes were determined by quantitative real-time PCR. Two cngc2 loss-of-function mutants were used: cngc2-3 and defence not death1 (which expresses cytosolic aequorin). Extracellular ATP-induced transient depolarization of Arabidopsis root elongation zone epidermal PM voltage was Ca²⁺ dependent, requiring CNGC2 but not CNGC4 (its channel co-subunit in immunity signalling). Activation of PM Ca²⁺ influx currents also required CNGC2. The eATP-induced [Ca²⁺ ]_cyt increase and transcriptional response in cngc2 roots were significantly impaired. CYCLIC NUCLEOTIDE-GATED CHANNEL2 is required for eATP-induced epidermal Ca²⁺ influx, causing depolarization leading to [Ca²⁺ ]_cyt increase and damage-related transcriptional response.

SUBMITTER: Wang L

PROVIDER: S-EPMC9211375 | biostudies-literature | 2022 Apr

REPOSITORIES: biostudies-literature

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Arabidopsis thaliana CYCLIC NUCLEOTIDE-GATED CHANNEL2 mediates extracellular ATP signal transduction in root epidermis.

Wang Limin L Ning Youzheng Y Sun Jian J Wilkins Katie A KA Matthus Elsa E McNelly Rose E RE Dark Adeeba A Rubio Lourdes L Moeder Wolfgang W Yoshioka Keiko K Véry Anne-Aliénor AA Stacey Gary G Leblanc-Fournier Nathalie N Legué Valérie V Moulia Bruno B Davies Julia M JM

The New phytologist 20220220 2

Damage can be signalled by extracellular ATP (eATP) using plasma membrane (PM) receptors to effect cytosolic free calcium ion ([Ca2+ ]cyt ) increase as a second messenger. The downstream PM Ca2+ channels remain enigmatic. Here, the Arabidopsis thaliana Ca2+ channel subunit CYCLIC NUCLEOTIDE-GATED CHANNEL2 (CNGC2) was identified as a critical component linking eATP receptors to downstream [Ca2+ ]cyt signalling in roots. Extracellul ...[more]

PMID: 35075689

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Project description:The cyclic nucleotide-gated channels, CNGC19 and CNGC20, are the sole members of the highly isolated evolutionary group IV-A in Arabidopsis plants. Prior studies have shown that the expression of both CNGC19 and CNGC20 genes are induced by salinity and biotic stress. In this report, CNGC19 and CNGC20 were determined to localize to the vacuolar membrane. Co-expression of CNGC19 and CNGC20 increased the efficiency of vacuolar localization. CNGC19 and CNGC20 are, therefore, vacuolar membrane channels that are hypothesized to mediate plant response to salinity and biotic stress. Plant cyclic nucleotide-gated channels (CNGCs) are implicated in the uptake of both essential and toxic cations, Ca2+ signalling, and responses to biotic and abiotic stress. The 20 CNGC paralogues of Arabidopsis are divided into five evolutionary groups. Group IV-A is highly isolated and consists only of two closely spaced genes, CNGC19 and CNGC20. Prior studies have shown that both genes are induced by salinity and biotic stress. A unique feature of CNGC19 and CNGC20 is their long hydrophilic N-termini. To determine the subcellular locations of CNGC19 and CNGC20, partial and full-length fusions to GFP(S65T) were generated. Translational fusions of the N-termini of CNGC19 (residues 1–171) and CNGC20 (residues 1–200) to GFP(S65T) were targeted to punctate structures when transiently expressed in leaf protoplasts. In the case of CNGC20, but not CNGC19, the punctate structures were co-labelled with a marker for the Golgi. The full-length CNGC19-GFP fusion co-localized with markers for the vacuole membrane (αTIP- and γTIP-mCherry). Vacuole membrane labelling by the full-length CNGC20-GFP fusion was also observed, but the signal was weak and accompanied by numerous punctate signals that did not co-localize with αTIP- or γTIP-mCherry. These punctate structures diminished, and localization of full-length CNGC20-GFP to the vacuole increased, when it was co-expressed with the full-length CNGC19-mCherry. Vacuole membrane labelling was also detected in planta via immunoelectron microscopy using a CNGC20-antiserum on cryopreserved ultrathin sections of roots. We hypothesize that the role of group IV-A CNGCs is to mediate the movement of cations between the central vacuole and the cytosol in response to certain types of abiotic and biotic stress.

Dataset Information

Arabidopsis thaliana CYCLIC NUCLEOTIDE-GATED CHANNEL2 mediates extracellular ATP signal transduction in root epidermis.

Publications

Arabidopsis thaliana CYCLIC NUCLEOTIDE-GATED CHANNEL2 mediates extracellular ATP signal transduction in root epidermis.

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