Rationale: Increasing evidence suggests that tumor necrosis factor (TNF)- plays a key role in pulmonary injury caused by environmental ozone (O₃) in animal models and human subjects. We previously determined that mice genetically deficient in TNF response are protected from lung inflammation and epithelial injury after O₃ exposure. Objectives: The present study was designed to determine the molecular mechanisms of TNF-receptor (TNF-R)-mediated lung injury induced by O₃. Methods: TNF-R knockout (Tnfr^-/-) and wild-type (Tnfr^+/+) mice were exposed to 0.3-ppm O₃ or air (6, 24, 48 hr), and lung RNA and proteins were prepared. Mice deficient in p50 nuclear factor (NF)-B (Nfkb1^-/-) or c-Jun-NH₂-terminal kinase 1 (Jnk1^-/-), and wild-type controls (Nfkb1^+/+, Jnk1+/+) were exposed to O₃ (48 hr), and the role of NF-B and mitogen activated protein kinase (MAPK) as downstream effectors of lung injury was analyzed by bronchoalveolar lavage analyses. Results: O₃-induced early activation of TNF-R-adaptor complex formation was attenuated in Tnfr^-/- mice compared with Tnfr^+/+ mice. O₃ significantly activated lung NF-B in Tnfr^+/+ mice before the development of lung injury. Basal and O₃-induced NF-B activity was suppressed in Tnfr^-/- mice. Compared with Tnfr^+/+ mice, MAPKs and activator protein-1 (AP-1) was lower in Tnfr^-/- mice basally and after O₃. Furthermore, inflammatory cytokines including MIP-2 were differentially expressed in Tnfr^-/- and Tnfr^+/+ mice after O₃. O₃-induced lung injury was significantly reduced in Nfkb1^-/- and Jnk1^-/- mice relative to respective controls. Conclusions: Results suggest that NF-B and MAPK/AP-1 signaling pathways are essential in TNF-R-mediated pulmonary toxicity induced by O₃.