Voltage-dependent anion channel of Arabidopsis hypocotyls: nucleotide regulation and pharmacological properties.

Abstract

Plasma membrane anion channels are thought to play important roles in osmoregulation and signal transduction in higher plant cells. Knowledge of their pharmacology and regulation is of importance to unravel their physiological functions. In this study, we explore the pharmacological properties and the nucleotide regulation of the voltage-dependent anion channel of Arabidopsis hypocotyls. The pharmacological profile of this channel is characterized by a low sensitivity to most anion channel blockers. It is inhibited by niflumic acid with an IC50 of 80 microM, but poorly sensitive to IAA-94 and NPPB and insensitive to 9-AC and DIDS. Nucleotides alter the amplitude, the kinetics and the voltage-dependence of the channel. The main effect of nucleotides is a shift of the voltage-dependent gate of the channel toward depolarized potentials leading to a strong reduction of the current amplitude. This regulation does not require ATP hydrolysis as nonhydrolyzable ATP analogues-AMPPNP and ATP gamma S-also regulate the anion current. This suggests that a nucleotide binding site is involved in the regulation. The study of the properties of this putative nucleotide binding site reveals that (i) ATP regulates the channel with an EC50 of 0.7 mM, (ii) adenyl nucleotides modulate the channel with the following order of effectiveness: ATP > ADP > > AMP, and (iii) thiophosphate nucleotide analogues are the most potent agonists with EC50 in the range of 80 microM. The hypothesis that this regulation may couple the electrical properties of the membrane with the metabolic status of the cell is discussed.