Root hairs proliferation and warming strongly influence exudate release, enzyme activities and microbial substrate utilization. However, how the presence of root hairs regulates those processes in the rhizosphere under elevated temperature is poorly known. To clarify these interactions, a wild type maize (with root hairs) and its hairless mutant were grown for 3 weeks at 20 and 30 °C, respectively. We combined zymography (localize hotspots of β-glucosidase) with substrate-induced respiration and microcalorimetry to monitor exudate effects on enzyme kinetics, microbial growth and heat production in the rhizosphere hotspots in response to warming. Root hairs effects were more pronounced at the elevated temperature: i) β-glucosidase activity of the wild type at 30 °C was 21% higher than that of the hairless maize; ii) temperature shifted the microbial growth strategy, whereas root hairs promoted the fraction of growing microbial biomass; iii) Km and the activation energy for β-glucosidase under the hairless mutant was lower than that under wild maize. These results suggest that microorganisms inhabiting hotspots of the wild type synthesized more enzymes to fulfill their higher energy and nutrient demands than those of the hairless mutant. In contrast, at higher temperature the hairless maize produced an enzyme pool with higher efficiencies rather than higher enzyme production, enabling metabolic needs to be met at lower cost. We therefore conclude that root hairs play an important role in regulating enzyme systems and microbial growth to adapt to climate warming. © 2020 Elsevier Ltd