AbstractThe leaf vascular bundle sheath cells (BSCs) that tightly envelop the leaf veins, are a selective and dynamic barrier to xylem-sap water and solutes radially entering the mesophyll cells. Under normal conditions, xylem-sap pH of <6 is presumably important for driving and regulating the transmembranal solute transport. Having discovered recently a differentially high expression of a BSCs proton pump, AHA2, we now test the hypothesis that it regulates this pH and leaf radial water fluxes.We monitored the xylem-sap pH in the veins of detached leaves of WT Arabidopsis, AHA mutants, and aha2 mutants complemented with AHA2 gene solely in BSCs. We tested an AHA inhibitor and stimulator, and different pH buffers. We monitored their impact on the xylem-sap pH and the whole leaf hydraulic conductance (Kleaf), and the effect of pH on the water osmotic permeability (Pf) of isolated BSCs protoplasts.Our results demonstrated that AHA2 is necessary for xylem-sap acidification, and in turn, for elevating Kleaf. Conversely, knocking out AHA2 alkalinized the xylem-sap. Also, elevating xylem sap pH to 7.5 reduced Kleaf and elevating external pH to 7.5 decreased the BSCs Pf.All these demonstrate a causative link between AHA2 activity in BSCs and leaf radial water conductance.One-sentence summaryBundle-sheath cells can control the leaf hydraulic conductance by proton-pump-regulated xylem sap pH