Patch-clamp analysis establishes a role for an auxin binding protein in the auxin stimulation of plasma membrane current in Zea mays protoplasts

Abstract

The electrical response of Zea mays protoplasts to different auxins and to antibodies raised against an ER-located auxin binding protein from maize (Zm-ERabp1), was investigated using the patch-clamp technique (whole-cell configuration). Following a lag-phase of 30-40 seconds, indole-3-acetic acid and 1-naphthylacetic acid induced an outwardly directed current of positive charge in a concentration-dependent manner. This current was further increased by the fungal toxin fusicoccin (FC). The current was observed only in the presence of Mg2+-ATP in the patch-pipette and was abolished after addition of erythrosin B, an inhibitor of H+-ATPase, to the protoplasts indicating that the plasma membrane H+-ATPase is activated by auxins and fusicoccin. Addition of antibodies directed against Zm-ERabp1 abolished the current induced by auxins, without affecting the response of protoplasts to fusicoccin. Antibodies directed against a peptide representing part of the putative auxin binding domain of Zm-ERabp1 showed auxin agonist activity, stimulating an outwardly directed membrane current in the absence of auxin. These results suggest that (i) Zm-ERabp1 or antigenically related proteins represent a site for auxin perception through which the plasma membrane H+-ATPase is activated, and (ii) that the activation of the H+-ATPase by such proteins is initiated from outside the plasma membrane.