Copper oxide nanoparticles (nano-CuO) are recognized as an emerging pollutant. Arbuscular mycorrhizal fungi (AMF) can mitigate the adverse impacts of various pollutants on host plants. However, AMF’s mechanism for alleviating nano-CuO phytotoxicity remains unclear. The goal of this study was to evaluate how AMF inoculations affect the physiological features of Canna indica seedlings exposed to nano-CuO stress. Compared with the non-AMF inoculated treatment, AMF inoculations noticeably improved plant biomass, mycorrhizal colonization, leaf chlorophyll contents, and the photosynthetic parameters of C. indica under nano-CuO treatments. Moreover, AMF inoculation was able to significantly mitigate nano-CuO stress by enhancing antioxidant enzyme activities and decreasing ROS levels in the leaves and roots of C. indica, thus increasing the expression of genes involved in the antioxidant response. In addition, AMF inoculation reduced the level of Cu in seedlings and was associated with an increased expression of Cu transport genes and metallothionein genes. Furthermore, AMF inoculations increased the expression levels of organic acid metabolism-associated genes while facilitating organic acid secretion, thus reducing the accumulation of Cu. The data demonstrate that AMF–plant symbiosis is a feasible biocontrol approach to remediate nano-CuO pollution.