Plant Science 154 2000 43 – 51
Molecular cloning of a novel water channel from rice: its products expression in Xenopus oocytes and involvement in chilling
tolerance
Le-gong Li
1
, Shi-fang Li
2
, Yan Tao, Yoshichika Kitagawa
Laboratory of Plant Genetic Engineering, Biotechnology Institute, Akita Prefectural Uni6ersity, Ogata, Akita
010
-
0444
, Japan Received 15 March 1999; received in revised form 28 October 1999; accepted 1 December 1999
Abstract
Water channel proteins, aquaporins, play a fundamental role in transmembrane water movements in plants. We isolated rice cDNA, rwc
1
, by screening a rice Oryza sati6a cv. Josaeng Tongil cDNA library using a conserved motif of aquaporins. Like other aquaporin genes, rwc
1
encodes a 290-residue protein with six putative transmembrane domains. The derived amino acid sequence of RWC1 shows high homology with PIP1 plasma membrane intrinsic protein 1 subfamily members, which suggest it
is localized in the plasma membrane. Injection of its cRNA into Xenopus oocytes increased the osmotic water permeability of the oocytes 2 – 3 times. Northern analysis showed that rice aquaporin genes are expressed in rice seedling leaves and roots, but that
it disappeared from the root 6 h after osmotic stress began and that the transcript level remained low for about 24 h, then recovered. The time course of rice aquaporin gene-expression under osmotic stress was correlated with time course of turgor
transition in plant. On the other hand, the levels of rice aquaporin gene-transcripts in leaves under chilling and recovery temperature depend on the pretreatment of mannitol for short time. This variation of the transcripts shown that rice aquaporin
genes may play an important role in response to water stress-induced chilling tolerance. © 2000 Elsevier Science Ireland Ltd. All rights reserved.
Keywords
:
Rice; Water channel; Water stress; Chilling tolerance; Xenopus oocyte www.elsevier.comlocateplantsci
1. Introduction
Transmembrane water flow is a fundamental process of life. Although water permeability as a
biophysical feature of cell membrane, the molecu- lar pathway of transmembrane water movement
remained unknown until the discovery of aquapor- ins [1]. The recent discovery that plants express
numerous aquaporins in both the plasma mem- brane and the tonoplast has changed our view of
how plant cells regulate transmembrane water movement [2,3]. Water channel proteins aquapor-
ins belong to the major intrinsic protein MIP superfamily that permit the passage of specific
molecules through biological membranes. Since the first aquaporin AQP1 was identified in hu-
man erythrocytes [1], many more have been iso- lated from various organisms, including bacteria,
plants, and animals [3]. In plants, many MIP genes have been isolated. They are encoded by several
gene families and are hydrophobic integral mem- brane proteins that range in apparent molecular
mass from 23 to 31 kDa [4]. Since g-tonoplast intrinsic protein g-TIP was first recognized as a
plant aquaporin [5], more different genes have
The nucleotide sequence data reported appeared in EMBL, Gen- Bank and DDBJ Nucleotide sequence Database under the accession
number AB009665. Corresponding author.
E-mail address
:
kitagawaagri.akita-pu.ac.jp Y. Kitagawa
1
Permanent address: Shanghai Institute of Plant Physiology, Chi- nese Academy of Sciences, Shanghai 200032, People’s Republic of
China.
2
Present address: Institute of Plant Protection, Chinese Academy of Agricultural Sciences, 2 West Yuanmingyuan Road, Beijing
100094, China. Le-gong Li and Shi-fang Li contributed equally to this work.
0168-945200 - see front matter © 2000 Elsevier Science Ireland Ltd. All rights reserved. PII: S 0 1 6 8 - 9 4 5 2 9 9 0 0 2 6 9 - 1
been identified [6]. Their products are located in the tonoplast and plasma membrane. Sequence
comparisons have shown a high homology be- tween plant aquaporins; all published sequences
are clearly of TIP or plasma membrane intrinsic protein PIP members. These relationships even
extend to the PIP1 and PIP2 subfamilies, origi- nally introduced by Kammerloher et al. [7]; easily
classified as PIP1 or PIP2 based on specific arrays of amino acids at the N- and C-termini.
Although DNA sequences with high homology to the aquaporin genes have been identified in
several plant species [4], the water permeability and function haven’t been determined, only a few
of the gene products have been characterized [6 – 11]. Their roles in response to various physiologi-
cal and stress conditions warrants further research. Here we report the isolation of the first water
channel gene to be identified in rice, the activity of its water permeability, and the possible role in
response to osmotic and chilling stress.
2. Materials and methods
