Lesion mimic mutants are used to elucidate mechanisms controlling plant responses to pathogen attacks and environmental stresses. Here, a new rice (Oryza sativa) lesion mimic mutant, natural blight leaf 3 (nbl3), was identified from T-DNA insertion lines. The causative gene, OsNBL3, encodes a mitochondria-localized pentatricopeptide repeat (PPR) protein. The nbl3 mutant exhibited spontaneous cell death and H₂O₂ accumulation as evidenced by Trypan blue, 3,3’-diaminobenzidine and tetranitroblue tetrazolium chloride analyses. Additionally, nbl3 displayed enhanced resistance to the fungal and bacterial pathogens Magnaporthe oryzae and Xanthomonas oryzae pv. oryzae. This resistance was consistent with the upregulation of several defence-related genes; thus defence responses were induced in nbl3. RNA interference lines of OsNBL3 exhibited enhanced disease resistance similar to that of nbl3, while the disease resistance of overexpression lines did not differ from that of the wild type. In addition, nbl3 displayed improved tolerance to salt treatment, accompanied by upregulation of several salt-associated marker genes. Disruption of OsNBL3 leads to destruction of mitochondria and elevated use of alternative respiratory pathways. OsNBL3 was found to mainly participate in the splicing of mitochondrial gene nad5 intron 4. Overall, the results demonstrated that a PPR protein was involved in disease resistance and salt tolerance in rice.