Improved understanding of the links between aboveground production and allocation of photosynthate to belowground processes and the temporal variation in those links is needed to interpret observations of belowground carbon cycling processes. Here, we show that combining a trenching manipulation with high-frequency soil respiration measurements in a temperate hardwood forest permitted identification of the temporally variable influence of roots on diel and seasonal patterns of soil respiration. The presence of roots in an untrenched plot caused larger daily amplitude and a 2–3 h delay in peak soil CO2 efflux relative to a root-free trenched plot. These effects cannot be explained by differences in soil temperature, and they were significant only when a canopy was present during the growing season. This experiment demonstrated that canopy processes affect soil CO2 efflux rates and patterns at hourly and seasonal time scales, and it provides evidence that root and microbial processes respond differently to environmental factors.