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Plant and Cell Physiology Advance Access originally published online on November 27, 2008
Plant and Cell Physiology 2009 50(1):151-160; doi:10.1093/pcp/pcn181
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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 email: journals.permissions@oxfordjournals.org

Acceleration of Vacuolar Regeneration and Cell Growth by Overexpression of an Aquaporin NtTIP1;1 in Tobacco BY-2 Cells

Emiko Okubo-Kurihara1, Toshio Sano1,2, Takumi Higaki1, Natsumaro Kutsuna1,2 and Seiichiro Hasezawa1,2,*

1Department of Integrated Biosciences, Graduate School of Frontier Sciences, The University of Tokyo, Kashiwanoha 5-1-5, Kashiwa, Chiba, 277-8562 Japan
2Institute for Bioinformatics Research and Development (BIRD), Japan Science and Technology Agency (JST), Japan

*Corresponding author: E-mail, hasezawa{at}k.u-tokyo.ac.jp.


  Abstract

Aquaporin is a water channel that increases water permeability through membranous structures. In plants, vacuoles are essential organelles that undergo dynamic volume changes during cell growth. To understand the contribution of aquaporins to plant cell growth, we developed a transgenic tobacco BY-2 cell line overexpressing the tonoplast intrinsic protein (TIP), {gamma}TIP. Vacuolar membranes of isolated vacuoles from {gamma}TIP-overexpressing cells showed higher water permeation activities than those from wild-type cells. We then examined the role of {gamma}TIP in vacuolar regeneration of evacuolated tobacco BY-2 protoplasts (miniprotoplasts). Vacuolar regeneration from thin to thick tube-network vacuoles and subsequent development of large vacuoles was accelerated in miniprotoplasts of this cell line. A parallel increase in the rate of cell expansion indicated a tight relationship between vacuolar development and cellular volume increases. Interestingly, overexpression of tobacco {gamma}TIP also enhanced cell division. Thus, increased vacuolar aquaporin activity may accelerate both cell expansion and cell division by increasing water permeability through the vacuolar membrane.

Keywords: {gamma}TIP - Vauolar regeneration - Cell growth - Aquaporin - Tobacco BY-2 cell - Miniprotoplast

Abbreviations: BY-2, Bright Yellow 2; BY-GV, BY-2 cells expressing GFP–AtVAM3 fusion proteins; BY-TIPG, BY-2 cells expressing NtTIP1;1–GFP fusion proteins; FM4-64, N-(3-triethylammoniumpropyl)-4-[6-(4-(diethylamino)phenyl)hexatrienyl]-pyridinium dibromide; GFP, green fluorescent protein; MIP, major intrinsic protein; Ng, Nicotiana glauca; Nt, Nicotiana tabacum; Pf, permeability coefficient; Pg, Panax ginseng; PM, plasma membrane; TIP, tonoplast intrinsic protein; vag, V-ATPase subunit G; VM, vacuolar membrane.

The nucleotide sequence of tobacco {gamma}TIP, NtTIP1;1, reported in this paper has been submitted in GenBank as accession number AB371711.

(Received October 29, 2008; Accepted November 19, 2008)
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