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JXB Advance Access published online on August 28, 2006
Journal of Experimental Botany, doi:10.1093/jxb/erl090
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© 2006 The Author(s). This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Received April 1, 2006
Accepted June 21, 2006

RESEARCH PAPER

Cloning of an H+-PPase gene from Thellungiella halophila and its heterologous expression to improve tobacco salt tolerance

Feng Gao 1, Qiang Gao 1, XiaoGuang Duan 1, GuiDong Yue 1, AiFang Yang 1 *, and JuRen Zhang 1

1 School of Life Science, Shandong University, Jinan 250100, PR China

* To whom correspondence should be addressed.
AiFang Yang, E-mail: nihaohua{at}263.net


  Abstract

An H+-pyrophosphatase (PPase) gene named TsVP involved in basic biochemical and physiological mechanisms was cloned from Thellungiella halophila. The deduced translation product has similar characteristics to H+-PPases from other species, such as Arabidopsis and rice, in terms of bioinformation. The heterologous expression of TsVP in the yeast mutant ena1 suppressed Na+ hypersensitivity and demonstrated the function of TsVP as an H+-PPase. Transgenic tobacco overexpressing TsVP had 60% greater dry weight than wild-type tobacco at 300 mM NaCl and higher viability of mesophyll protoplasts under salt shock stress conditions. TsVP and AVP1, another H+-PPase from Arabidopsis, were heterologously expressed separately in both the yeast mutant ena1 and tobacco. The salt tolerance of TsVP or AVP1 yeast transformants and transgenic tobacco were improved to almost the same level. The TsVP transgenic tobacco lines TL3 and TL5 with the highest H+-PPase hydrolytic activity were studied further. These transgenic tobacco plants accumulated 25% more solutes than wild-type plants without NaCl stress and 20-32% more Na+ under salt stress conditions. Although transgenic tobacco lines TL3 and TL5 accumulated more Na+ in leaf tissues, the malondialdehyde content and cell membrane damage were less than those of the wild type under salt stress conditions. Presumably, compartmentalization of Na+ in vacuoles reduces its toxic effects on plant cells. This result supports the hypothesis that overexpression of H+-PPase causes the accumulation of Na+ in vacuoles instead of in the cytoplasm and avoids the toxicity of excessive Na+ in plant cells.

Keywords: Arabidopsis thaliana; H+-PPase; salt tolerance; Thellungiella halophila.
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