Coumarin enhances nitrate uptake in maize roots through modulation of plasma membrane H⁺ -ATPase activity

Coumarin is one of the simplest plant secondary metabolites, widely distributed in the plant kingdom, affecting root form and function, including anatomy, morphology and nutrient uptake. Although, some plant responses to coumarin have been described, comprehensive knowledge of the physiological and molecular mechanisms is lacking. Maize seedlings exposed to different coumarin concentrations, alone or in combination with 200 μm nitrate (NO₃^- ), were analysed, through a physiological and molecular approach, to elucidate action of coumarin on net NO₃^- uptake rate (NNUR). In detail, the time course of NNUR, plasma membrane (PM) H⁺ -ATPase activity, proton pumping and related gene expression (ZmNPF6.3, ZmNRT2.1, ZmNAR2.1, ZmHA3 and ZmHA4) were evaluated. Coumarin alone did not affect nitrate uptake, PM H⁺ -ATPase activity or transcript levels of ZmNRT2.1 and ZmHA3. In contrast, coumarin alone increased ZmNPF6.3, ZmNAR2.1 and ZmHA4 expression in response to abiotic stress. When coumarin and NO₃^- were concurrently added to the nutrient solution, a significant increase in the NNUR, PM H⁺ -ATPase activity, together with ZmNAR2.1:ZmNRT2.1 and ZmHA4 expression was observed, suggesting that coumarin affected the inducible component of the high affinity transport system (iHATS), and this effect appeared to be mediated by nitrate. Moreover, results with vanadate, an inhibitor of the PM H⁺ -ATPase, suggested that this enzyme could be the main target of coumarin. Surprisingly, coumarin did not affect PM H⁺ -ATPase activity by direct contact with plasma membrane vesicles isolated from maize roots, indicating its possible elicitor role in gene transcription.