Further insights into underlying mechanisms for the release of biological nitrification inhibitors from sorghum roots

Background Sorghum roots release two categories of biological nitrification inhibitors (BNIs) – hydrophilic-BNIs and hydrophobic-BNIs. Earlier research indicated that rhizosphere pH and plasma membrane (PM) H+ATPase are functionally linked with the release of hydrophilic BNIs, but the underlying mechanisms are not fully elucidated. This study is designed to reveal further insights into the regulatory mechanisms of BNIs release in root systems, using three sorghum genetic stocks. Methods Sorghum plants were grown in a hydroponic system with pH of nutrient solutions ranging from 3.0 ̴ 9.0. Pharmacological agents [(fusicoccin and vanadate) and anion-channel blockers (−niflumic acid (NIF) and anthracene-9-carboxylate (A9C)] were applied to root exudate collection solutions; BNI activity was determined with luminescent Nitrosomonas europaea bioassay. Sorgoleone levels in root exudates and H+ excretion from roots were determined. Two-phase partitioning system is used to isolate root plasma membrane (PM) and H+ ATPase activity was determined. Results A decrease in rhizosphere pH improved the release of hydrophilic-BNIs from roots of all the three sorghum genotypes, but had no effect on the release of hydrophobic-BNIs. Hydrophobic-BNI activity and sorgoleone levels in root-DCM wash are positively correlated. Fusicoccin promoted H+extrusion and stimulated the release of hydrophilic-BNIs. Vanadate, in contrast, suppressed H+ extrusion and lowered the release of hydrophilic-BNIs. Anion-channel blockers did not inhibit the release of hydrophilic BNIs, but enhanced H+-extrusion and hydrophilic-BNIs release. Conclusion Rhizosphere pH has a major influence on hydrophilic-BNIs release, but not on the release of hydrophobic-BNIs. The low rhizosphere pH stimulated PM-H+ ATPase activity; H+-extrusion is closely coupled with hydrophilic-BNIs release. Anion-channel blockers stimulated H+ extrusion and hydrophilic-BNIs release. Our results indicate that some unknown membrane transporters are operating the release of protonated BNIs, which may compensate for charge balance when transport of other anions is suppressed using anion-channel blockers. A new hypothesis is proposed for the release of hydrophilic-BNIs from sorghum roots.