Cucumber (Cucumis sativus) is one of the most widely cultivated vegetable crops in the world, and its yield is often reduced due to the infection of Botrytis cinerea (B. cinerea), which causes a serious disease. However, few genes involved in the response to B. cinerea have been identified in cucumber. In this study, we identified that CsWRKY10 plays a key role in the cucumber resistance to B. cinerea because that the overexpression of CsWRKY10 significantly increased the susceptibility to B. cinerea in cucumber. After the pathogen infection, the enzyme activities of catalase, superoxide dismutase and peroxidase in transgenic plants were affected, resulting in the decrease in reactive oxygen species (ROS) contents. In addition, the light microscopic images showed that overexpression of CsWRKY10 promoted the spore germination and mycelia elongation of B. cinerea in cucumber. Importantly, after B. cinerea infection, the contents of jasmonic acid (JA) are decreased, and the expression levels of JA- and salicylic acid- related defence genes significantly changed in transgenic plants. In contrast, overexpression of CsWRKY10 enhanced resistance to Corynespora cassiicola in cucumber. Collectively, this study indicated that CsWRKY10 negatively regulates the resistance of cucumber to B. cinerea by reducing the ROS contents and inhibiting the JA-mediated resistance signalling pathway, but strengthens resistance to Corynespora cassiicola.