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Eukaryotic Cell, September 2009, p. 1439-1448, Vol. 8, No. 9
1535-9778/09/$08.00+0     doi:10.1128/EC.00059-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

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

Miroslav Zivic,1* Marko Popovic,2 Natasa Todorovic,3 and Zeljko Vucinic2

Department of Bio-Chemical and Medical Sciences, State University of Novi Pazar, Vuka Karadzica bb, 36300 Novi Pazar, Serbia,1 Institute for Multidisciplinary Research, Kneza Viseslava 1, 11030 Belgrade, Serbia,2 Institute for Biological Research Sinisa Stankovic, Despota Stefana 142, 11000 Belgrade, Serbia3

Received 21 February 2009/ Accepted 30 June 2009

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– ≥ NO3– > Br– > Cl– > SO42– = 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 µM), 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.

* Corresponding author. Mailing address: Department of Bio-Chemical and Medical Sciences, State University of Novi Pazar, Vuka Karadzica bb, 36300 Novi Pazar, Serbia. Phone: 381 11 2078308. Fax: 381 11 2761433. E-mail: mzivic{at}np.ac.yu

{triangledown} Published ahead of print on 10 July 2009.

Eukaryotic Cell, September 2009, p. 1439-1448, Vol. 8, No. 9
1535-9778/09/$08.00+0     doi:10.1128/EC.00059-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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