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.