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Expression of AoNHX1 increases salt tolerance of rice and Arabidopsis, and bHLH transcription factors regulate AtNHX1 and AtNHX6 in Arabidopsis

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Expression of AoNHX1 from the mangrove Avicennia increases salt tolerance of rice and Arabidopsis, and specific bHLH transcription factors regulate AtNHX1 and AtNHX6 in Arabidopsis to mediate the salinity response.

Abstract

Improving crop plants to better tolerate soil salinity is a challenging task. Mangrove trees such as Avicennia officinalis have special adaptations to thrive in high salt conditions, which include subcellular compartmentalization of ions facilitated by specialized ion transporters. We identified and characterized two genes encoding Na+/H+ exchangers AoNHX1 and AoNHX6 from Avicennia. AoNHX1 was present in the tonoplast, while, AoNHX6 was localized to the ER and Golgi. Both NHXs were induced by NaCl treatment, with AoNHX1 showing high expression levels in the leaves and AoNHX6 in the seedling roots. Yeast deletion mutants (ena1-5Δ nha1Δ nhx1Δ and ena1-5Δ nha1Δ vnx1Δ) complemented with AoNHX1 and AoNHX6 showed increased tolerance to both NaCl and KCl. Expression of AoNHX1 and AoNHX6 in the corresponding Arabidopsis mutants conferred enhanced NaCl tolerance. The underlying molecular regulatory mechanism was investigated using AtNHX1 and AtNHX6 in Arabidopsis. We identified two basic helix–loop–helix (bHLH) transcription factors AtMYC2 and AtbHLH122 as the ABA-mediated upstream regulators of AtNHX1 and AtNHX6 by chromatin immunoprecipitation. Furthermore, expression of AtNHX1 and AtNHX6 transcripts was reduced in the atmyc2 and atbhlh122 mutants. Lastly, transgenic rice seedlings harboring pUBI::AoNHX1 showed enhanced salt tolerance, suggesting that this gene can be exploited for developing salt-tolerant crops.

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Acknowledgements

This research grant was supported by the Singapore National Research Foundation under its Environment and Water Research Programme and administered by PUB, Singapore’s National Water Agency, Singapore, NRF-EWI-IRIS (R-706-000-010-272 and R-706-000-040-279). We thank the NParks Singapore for granting us permission to collect the mangrove samples from Berlayer Creek and Sungei Buloh Wetland Reserves (NP/RP 12-002-1 & NP/RP 12-002-2). Our sincere thanks to Dr. Hana Sychrová for providing the yeast strains used in this study.

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  1. Department of Biological Sciences, National University of Singapore, 14 Science Drive 4, Singapore, 117543, Singapore

    Pannaga Krishnamurthy, Bhushan Vishal, Kaijie Khoo, Sivamathini Rajappa, Chiang-Shiong Loh & Prakash P. Kumar

  2. NUS Environmental Research Institute (NERI), National University of Singapore, #02-01, T-Lab Building, 5A Engineering Drive 1, Singapore, 117411, Singapore

    Pannaga Krishnamurthy, Chiang-Shiong Loh & Prakash P. Kumar

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  1. Pannaga Krishnamurthy
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PK, LCS and PPK conceived the research plans; PK, KK and SR performed most of the experiments; BV carried out rice experiments; PK designed the experiments, analyzed the data and wrote the article with contributions of all the authors.

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Correspondence to Prakash P. Kumar.

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Communicated by Prakash Lakshmanan.

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Krishnamurthy, P., Vishal, B., Khoo, K. et al. Expression of AoNHX1 increases salt tolerance of rice and Arabidopsis, and bHLH transcription factors regulate AtNHX1 and AtNHX6 in Arabidopsis. Plant Cell Rep 38, 1299–1315 (2019). https://doi.org/10.1007/s00299-019-02450-w

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