RNA interference, mediated by small interfering RNAs (siRNAs), is a conserved regulatory process that has evolved as an antiviral defense mechanism in plants and animals. It is not known whether host cells also use siRNAs as an antibacterial defense mechanism in eukaryotes. Here, we report the discovery of an endogenous siRNA, nat-siRNAATGB2, that is specifically induced by the bacterial pathogen Pseudomonas syringae carrying effector avrRpt2. We demonstrate that the biogenesis of this siRNA requires DCL1, HYL1, HEN1, RDR6, NRPD1A, and SGS3. Its induction also depends on the cognate host disease resistance gene RPS2 and the NDR1 gene that is required for RPS2-specified resistance. This siRNA contributes to RPS2-mediated race-specific disease resistance by repressing PPRL, a putative negative regulator of the RPS2 resistance pathway.