Forest canopies maintain a high proportion of arthropod diversity. The drivers that structure these communities, however, are poorly understood. Therefore, integrative research connecting tree species identity and environmental stand properties with taxonomic and functional community composition of canopy arthropods is required. In this study, we investigated how the taxonomic, functional and trophic composition of arboreal spider communities is affected by tree species composition and associated differences in canopy structure and prey availability in temperate forests. We sampled canopy spiders as well as their potential prey using insecticidal fogging in monospecific and mixed stands of native European beech, native Norway spruce and non-native Douglas fir. Trophic metrics were obtained from stable isotope analysis and structural canopy properties were assessed with mobile laser scanning. Monospecific native spruce stands promoted local canopy spider abundance and diversity, but native beech and beech–conifer mixtures had the highest diversity at landscape scale. Spider community composition differed between monospecific stands, with broadleaf–conifer mixtures mitigating these differences. Irrespective of tree species identity, spider abundance, taxonomic diversity, functional richness and isotopic richness increased in structurally heterogeneous canopies with high prey abundances, but functional evenness and trophic divergence decreased. Our study shows that canopy spiders are differentially affected by tree species identity, canopy structure and prey availability. Broadleaf–conifer mixtures mitigated negative effects of (non-native) conifers, but positive mixture effects were only evident at the landscape scale. Structurally heterogeneous canopies promoted the dominance of only specific trait clusters. This indicates that intermediate heterogeneity might result in high stability of ecological communities.