Purinergic signaling activated by extracellular nucleotides and their derivative nucleosides trigger sophisticated signaling networks. The outcome of these pathways determine the capacity of the organism to survive under challenging conditions. Both extracellular ATP (eATP) and Adenosine (eAdo) act as primary messengers in mammals, essential for immunosuppressive responses. Despite the clear role of eATP as a plant damage-associated molecular pattern, the function of its nucleoside, eAdo, and of the eAdo/eATP balance in plant stress response remain to be fully elucidated. This is particularly relevant in the context of plant-microbe interaction, where the intruder manipulates the extracellular matrix. Here, we identify Ado as a main molecule secreted by the vascular fungus Fusarium oxysporum. We show that eAdo modulates the plant's susceptibility to fungal colonization by altering the eATP-mediated apoplastic pH homeostasis, an essential physiological player during the infection of this pathogen. Our work indicates that plant pathogens actively imbalance the apoplastic eAdo/eATP levels as a virulence mechanism.

To fire action-potential-like electrical signals, the vacuole membrane requires the two-pore channel TPC1, formerly called SV channel. The TPC1/SV channel functions as a depolarization-stimulated, non-selective cation channel that is inhibited by luminal Ca²⁺. In our search for species-dependent functional TPC1 channel variants with different luminal Ca²⁺ sensitivity, we found in total three acidic residues present in Ca²⁺ sensor sites 2 and 3 of the Ca²⁺-sensitive AtTPC1 channel from Arabidopsis thaliana that were neutral in its Vicia faba ortholog and also in those of many other Fabaceae. When expressed in the Arabidopsis AtTPC1-loss-of-function background, wild-type VfTPC1 was hypersensitive to vacuole depolarization and only weakly sensitive to blocking luminal Ca²⁺. When AtTPC1 was mutated for these VfTPC1-homologous polymorphic residues, two neutral substitutions in Ca²⁺ sensor site 3 alone were already sufficient for the Arabidopsis At-VfTPC1 channel mutant to gain VfTPC1-like voltage and luminal Ca²⁺ sensitivity that together rendered vacuoles hyperexcitable. Thus, natural TPC1 channel variants exist in plant families which may fine-tune vacuole excitability and adapt it to environmental settings of the particular ecological niche.