Plants have evolved sophisticated mechanisms to detect various forms of danger. Damage-associated molecular patterns (DAMPs) are endogenous danger molecules that are released from damaged cells and activate the innate immunity. Recent evidence suggests that plant extracellular self-DNA (esDNA) can serve as a DAMP molecule. However, the mechanisms by which esDNA functions are largely unknown. In this study, we confirmed that esDNA inhibits root growth and triggers reactive oxygen species (ROS) production in a concentration- and species-specific manner in Arabidopsis (Arabidopsis thaliana) and tomato (Solanum lycopersicum L.). Furthermore, by combining RNA sequencing, hormone measurement and genetic analysis, we found that esDNA-mediated growth inhibition and ROS production are achieved through the jasmonic acid (JA) signaling pathway. Specifically, esDNA induces JA production and the expression of JA responsive genes. The esDNA-mediated growth inhibition, ROS production and gene expression are impaired in the JA-related mutants. Finally, we found that the JA signaling pathway is required for the esDNA-elicited resistance against the pathogens Botrytis cinerea and Pseudomonas syringae pv. tomato DC3000. This finding highlights the importance of JA signaling in esDNA-mediated biological effects, thereby providing insight into how esDNA functions as a DAMP.