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JXB Advance Access published online on March 26, 2004
Journal of Experimental Botany, doi:10.1093/jxb/erh120
© 2004 by Oxford University Press
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Received August 1, 2003; accepted February 2, 2004
© 2004 Society for Experimental Biology

RESEARCH PAPER

Pulsing Cl- channels in coat cells of developing bean seeds linked to hypo-osmotic turgor regulation

Wen-Hao Zhang 1*, N. Alan Walker 2, John W. Patrick 3, and Stephen D. Tyerman 1

1 Department of Horticulture, Viticulture and Oenology, The University of Adelaide, Waite Campus PMB 1, Glen Osmond, SA 5064, Australia
2 Biophysics Department, School of Physics, The University of NSW, Kensington, NSW, 2052, Australia
3 School of Life and Environmental Sciences, The University of Newcastle, Newcastle, NSW 2308, Australia

* To whom correspondence should be addressed. E-mail: wen-hao.zhang{at}adelaide.edu.au.


  Abstract

Seed coat cells in the developing seeds of grain legumes release nutrients to the developing embryo. This occurs into an apoplastic space that separates the maternal (seed coat) and filial (embryo) generations. Protoplasts of seed coat cells from coats of Phaseolus vulgaris L. seeds were isolated and whole-cell current across their plasma membranes was characterized using the patch-clamp technique. A pulsing inward current that displayed a spontaneous activation and voltage-dependent inactivation was observed. The frequency and magnitude of the current pulses were positively dependent on cytoplasmic Cl- concentrations and independent of external cations. The pulse current was inhibited by DIDS and La3+, but not by Gd3+. Single channel events (conductance=18 pS) could be identified with the inactivating phase of the pulses. Together, these findings are consistent with the current being carried by a burst of Cl- efflux through Cl--permeable channels that activate almost simultaneously. Neomycin caused a reversible inhibition of the pulsed current, suggesting that its activation is likely to be modulated by an IP3-dependent intracellular Ca2+ release. The pharmacological profiles of Cl- efflux from excised seed coats were comparable with those of the Cl- channels in the whole cell configuration, suggesting that the Cl- channels may underpin Cl- efflux from the seed coats. Efflux of Cl- from the seed coats was also stimulated by hypo-osmotic treatment as was the frequency and magnitude of Cl- channel in whole-cell patch clamp experiments. This implies that the Cl- channels responsible for the pulsed Cl- currents are likely to be a component of the turgor-regulatory mechanism in developing bean seeds.

Key words: Hypo-osmotic treatment, neomycin, patch-clamp, Phaseolus vulgaris L., pulsed Cl- channels, seed coat.


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