Extracellular fragmented self-DNA (eDNA) is involved in plant responses to biotic stress

A growing body of evidence indicates that extracellular fragmented self-DNA (eDNA), by acting as a signaling molecule (or second messenger), triggers inhibitory effects on conspecific plants and functions as a damage-associated molecular pattern (DAMP). To evaluate early and late events in DAMP-dependent responses to eDNA, we extracted, fragmented and applied the tomato (Solanum lycopersicum) eDNA to tomato leaves. Non sonicated, intact self-DNA (intact DNA) was used as control. Early event analyses included the evaluation of plasma transmembrane potentials (Vm), cytosolic calcium variations (Ca2+cyt), the activity and subcellular localization of both voltage-gated and ligand-gated rectified K+ channels and the reactive oxygen species (ROS) subcellular localization and quantification. Late events included RNA-Seq transcriptomic analysis and qPCR validation of gene expression of tomato leaves exposed to tomato eDNA. eDNA induced a concentration-dependent Vm depolarization which was correlated to an increase in (Ca2+)cyt; this event induced the opening of K+ channels, with particular action on ligand-gated rectified K+ channels. Both eDNA-dependent (Ca2+)cyt and K+ increases were correlated to ROS production. In contrast, intact DNA produced no effects. eDNA modulated the expression of genes involved in plant-biotic interactions including pathogenesis-related proteins (PRP), calcium-dependent protein kinases (CPK1), heat shock transcription factors (Hsf), heat shock proteins (Hsp), receptor-like protein kinases (RLK) and ethylene-responsive factors (ERF). Several genes involved in calcium signaling, ROS scavenging and ion homeostasis were also modulated by eDNA. A comparison of tomato eDNA early and late responses with biotic stress caused by pathogens and herbivores indicated the occurrence of common DAMPs. Our results support the intriguing hypothesis that most of the plant reactions to pathogens and herbivores might be due to self DNA degradation as a consequence of the plant cell disruption/apoptosis. Fragmented DNA would then become an important and powerful second messenger able to trigger early and late responses to biotic stress.