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Plant and Cell Physiology Advance Access published online on May 1, 2008
Plant and Cell Physiology, doi:10.1093/pcp/pcn068
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© The Author 2008. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For Permissions, please e-mail: journals.permissions@oxfordjournals.org

Cryobehavior of the plasma membrane in protoplasts isolated from cold-acclimated Arabidopsis leaves is related to surface area regulation

Tomokazu Yamazaki, Yukio Kawamura and Matsuo Uemura

Institution addresses: The Iwate University 21st Century COE program (T.Y., Y.K., M.U.) and Cryobiofrontier Research Center, Faculty of Agriculture (M.U.), Iwate University, 3-18-8 Ueda, Morioka, Iwate 020-8550, Japan

Corresponding author: Yukio Kawamura, Address: Iwate University, 3-18-8 Ueda, Morioka, Iwate 020-8550, Japan. Tel: +81-19-621-6200, FAX: +81-19-621-6200, e-mail: ykawa{at}iwate-u.ac.jp


  Abstract

Extracellular freezing in plants results in dehydration and mechanical stresses upon the plasma membrane. Plants that acquire enhanced freezing tolerance after cold acclimation can withstand these two physical stresses. To understand the tolerance to freeze-induced physical stresses, the cryobehavior of the plasma membrane was observed using protoplasts isolated from cold-acclimated Arabidopsis thaliana leaves with the combination of a lipophilic fluorescent dye FM 1-43 and cryomicroscopy. We found that many vesicular structures appeared in the cytoplasmic region near the plasma membrane just after extracellular freezing occurred. These structures, referred to as freeze-induced vesicular structures (FIVs), then developed horizontally near the plasma membrane during freezing. There was a strong correlation between the increase in individual FIV size and the decrease in the surface area of the protoplasts during freezing. Some FIVs fused with their neighbors as the temperature decreased. Occasionally, FIVs fused with the plasma membrane, which may be necessary to relax the stress upon the plasma membrane during freezing. Vesicular structures resembling FIVs were also induced when protoplasts were mechanically pressed between a coverslip and slide glass. Fewer FIVs formed when protoplasts were subjected to hyperosmotic solution, suggesting that FIV formation is associated with mechanical stress rather than dehydration. Collectively, these results suggest that cold-acclimated plant cells may balance membrane tension in the plasma membrane by regulating surface area. This enables plant cells to withstand the direct mechanical stress imposed by extracellular freezing.

Keywords: cold acclimation - freezing tolerance - plasma membrane - surface area regulation

(Received March 18, 2008; Accepted April 24, 2008)
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