Isoprene emission from plants subject to a combination of ozone (O3) and nitrogen (N) has never been investigated. Cathay poplar (Populus cathayana) saplings were exposed to O3 (CF, charcoal-filtered air, NF, non-filtered ambient air and E-O3, non-filtered air +40ppb) and N treatments (N0, 0kgNha−1year−1, N50, 50kgNha−1year−1 and N100, 100kgNha−1year−1) for 96days. Increasing O3 exposure decreased isoprene emission (11.5% in NF and 57.9% in E-O3), as well as light-saturated photosynthetic rate (Asat) and chlorophyll content, while N load increased isoprene emission (19.6% in N50 and 33.4% in N100) as well as Asat and chlorophyll content. Although O3 and N interacted significantly in Asat, N did not mitigate the negative effects of O3 on isoprene emission, i.e. the combined effects were additive and did not interact. These results warrant more research on the combined effects of co-existing global change factors on future isoprene emission and atmospheric chemical processes.