To examine the potential roles of melatonin in Cd uptake, accumulation and detoxification in Malus plants, we exposed two different apple rootstocks varying greatly in Cd uptake and 2 accumulation to either 0 or 30 μM Cd together with 0 or 100 μM melatonin. Cd stress stimulated endogenous melatonin production to a greater extent in the Cd-tolerant M. baccata than in the Cd-susceptible M. micromalus "qingzhoulinqin". Melatonin application attenuated Cd-induced reductions in growth, photosynthesis, and enzyme activity, as well as ROS and MDA accumulation. Melatonin treatment more effectively restored photosynthesis, photosynthetic pigments, and biomass in Cd-challenged M. micromalus "qingzhoulinqin" than in Cd-stressed M. baccata. Exogenous melatonin lowered root Cd 2+ uptake, reduced leaf Cd accumulation, decreased Cd translocation factors (T f s), and increased root, stem, and leaf melatonin contents in both Cd-exposed rootstocks. Melatonin application increased both antioxidant concentrations and enzyme activities to scavenge Cd-induced ROS. Exogenous melatonin treatment altered the mRNA levels of several genes regulating Cd uptake, transport, and detoxification including HA7, NRAMP1, NRAMP3, HMA4, PCR2, NAS1, MT2, ABCC1, and MHX. Taken together, these results suggest that exogenous melatonin reduced aerial parts Cd accumulation and mitigated Cd toxicity in Malus plants, probably due to the melatonin-mediated Cd allocation in tissues, and induction of antioxidant defense system and transcriptionally regulated key genes involved in detoxification.