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Molecular and functional analyses of rice NHX-type Na+/H+ antiporter genes

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Abstract

We previously cloned a vacuolar Na+/H+ antiporter gene (OsNHX1) from rice (Oryza sativa). Here we identified four additional NHX-type antiporter genes in rice (OsNHX2 through OsNHX5) and performed molecular and functional analyses of those genes. The exon–intron structure of the OsNHX genes and the phylogenetic tree of the OsNHX proteins suggest that the OsNHX proteins are categorized into two subgroups (OsNHX1 through OsNHX4 and OsNHX5). OsNHX1, OsNHX2, OsNHX3, and OsNHX5 can suppress the Na+, Li+, and hygromycin sensitivity of yeast nhx1 mutants and their sensitivity to a high K+ concentration. The expression of OsNHX1, OsNHX2, OsNHX3, and OsNHX5 is regulated differently in rice tissues and is increased by salt stress, hyperosmotic stress, and ABA. When we studied the expression of β-glucuronidase (GUS) driven by either the OsNHX1 or the OsNHX5 promoter, we observed activity in the stele, the emerging part of lateral roots, the vascular bundle, the water pore, and the basal part of seedling shoots with both promoters. In addition, each promoter had a unique expression pattern. OsNHX1 promoter–GUS activity only was localized to the guard cells and trichome, whereas OsNHX5 promoter–GUS activity only was localized to the root tip and pollen grains. Our results suggest that the members of this gene family play important roles in the compartmentalization into vacuoles of the Na+ and K+ that accumulate in the cytoplasm and that the differential regulation of antiporter gene expression in different rice tissues may be an important factor determining salt tolerance in rice.

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Abbreviations

ABA:

Abscisic acid

ABRE:

ABA-responsive element

bZIP:

Basic-domain leucine zipper

EST:

Expressed sequence tag

GUS:

β-Glucuronidase

kb:

Kilobase

4-MU:

4-Methylumbelliferone

4-MUG:

4-Methylumbelliferyl-β-d-glucuronide

PAC:

P1-derived artificial chromosome

RACE:

Rapid amplification of cDNA ends

RGRP:

Rice Genome Research Program

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Acknowledgments

We thank Dr. Rajini Rao (Johns Hopkins University, USA) for kindly providing the yeast strains, K601 and R100. We also thank Drs. I. Mitsuhara and Y. Ohashi for kindly providing the vectors, pTN2 and pE2113–GUS. The cDNA clones including OsNHX2 and OsNHX5 and PAC clones including OsNHX1 and OsNHX5 were gifts from the Rice Genome Research Program. We thank Drs. Y. Nagamura and M. Yano for technical advice. We also thank C. Tsuiki, K. Toyoshima, T. Kataoka, and S. Li for technical assistance. This work was supported by Grants-in-Aid from the Ministry of Agriculture, Forestry and Fisheries of Japan (Development of Innovative Transgenic Plants no.2113 and Rice Genome Project MP-2126).

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Author notes

  1. Atsuko Nakamura

    Present address: Graduate School of Life and Environmental Sciences, Tsukuba University, Tennoudai 1-1-1, Tsukuba, Ibaraki, 305-8572, Japan

Authors and Affiliations

  1. Division of Plant Sciences, National Institute of Agrobiological Sciences, Kannondai 2-1-2, Tsukuba, Ibaraki, 305-8602, Japan

    Atsunori Fukuda, Naho Hara, Seiichi Toki & Yoshiyuki Tanaka

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  1. Atsunori Fukuda
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  2. Atsuko Nakamura
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  5. Yoshiyuki Tanaka
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Correspondence to Atsunori Fukuda.

Additional information

The nucleotide sequences reported in this paper have been submitted to the DDBJ/EMBL/GenBank with accession numbers AB531435 (OsNHX2), AB531433 (OsNHX3), and AB531434 (OsNHX5).

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Fukuda, A., Nakamura, A., Hara, N. et al. Molecular and functional analyses of rice NHX-type Na+/H+ antiporter genes. Planta 233, 175–188 (2011). https://doi.org/10.1007/s00425-010-1289-4

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