Outwardly Rectifying Anionic Channel from the Plasma Membrane of the Fungus Phycomyces blakesleeanus

Abstract

In the present report, by using a patch clamp technique, we provide, to our knowledge, the first detailed description of an anionic channel from filamentous fungi. The characterized channel, an outwardly rectifying anionic channel (ORAC), is the most prominent feature of the cell membrane of the fungus Phycomyces blakesleeanus in the absence of energizing substrates. The unitary conductance of the channel is 11.3 +/- 0.4 pS. It is characterized by a strong voltage dependence of the open-channel probability (z delta; the gating charge is 2.1 +/- 0.1), and the channel is activated by depolarization. The values of the time constants for voltage-induced activation and deactivation of 28 +/- 3 ms for tau(a) and 39 +/- 9 ms for tau(d) show that the ORAC is characterized by fast activation/deactivation kinetics. The ORAC shows strong selectivity for anions over cations and weak selectivity among anions, with a selectivity sequence of I- >= NO(3)(-) > Br(-) > Cl(-) > SO(4)(2-) = 4.8 > 4.4 > 2.2 > 1 > 0.55, which corresponds to Eisenman series 1. The channel is characterized by two open and two closed states, with dominant long open (tau(o2) = 35.0 +/- 3.9 ms) and long closed (tau(c2) = 166 +/- 28 ms) states occupying 63% +/- 8% and 79% +/- 3% of total open and closed times, respectively. The ORAC is insensitive to anthracene-9-carboxylic acid (< 200 mu M), but 2 mM malate reversibly inhibits 59% +/- 12% of the channel activity. Based on the electrophysiological properties of the channel, we propose that the ORAC plays a role in anion accumulation and in membrane potential regulation through local membrane depolarization.