The ELO family is involved in synthesizing very-long-chain fatty acids (VLCFAs) and VLCFAs play a crucial role in plant development, protein transport, and disease resistance, but the physiological function of the plant ELO family is largely unknown. Further, while nitric oxide synthase (NOS)-like activity acts in various plant environmental responses by modulating nitric oxide (NO) accumulation, how the NOS-like activity is regulated in such different stress responses remains misty. Here, we report that the yeast mutant Δelo3is defective in H2O2-triggered cell apoptosis with decreased NOS-like activity and NO accumulation, while its Arabidopsis homologous geneELO2 (ELO HOMOLOG 2)could complement such defects in Δelo3. The expression of this gene is enhanced and required in plant osmotic stress response because the T-DNA insertion mutantelo2is more sensitive to the stress than wild-type plants, andELO2expression could rescue the sensitivity phenotype ofelo2. In addition, osmotic stress-promoted NOS-like activity and NO accumulation are significantly repressed inelo2, while exogenous application of NO donors can rescue this sensitivity ofelo2in terms of germination rate, fresh weight, chlorophyll content, and ion leakage. Furthermore, stress-responsive gene expression, proline accumulation, and catalase activity are also repressed inelo2compared with the wild type under osmotic stress. In conclusion, our study identifies ELO2 as a pivotal factor involved in plant osmotic stress response and reveals its role in regulating NOS-like activity and NO accumulation.