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Molecular cloning and functional characterization of a putative sulfite oxidase (SO) ortholog from Nicotiana benthamiana

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Abstract

Sulfite oxidase (SO) catalyzes the oxidation of sulfite to sulfate and thus has important roles in diverse metabolic processes. However, systematic molecular and functional investigations on the putative SO from tobacco (Nicotiana benthamiana) have hitherto not been reported. In this work, a full-length cDNA encoding putative sulfite oxidase from N. benthamiana (NbSO) was isolated. The deduced NbSO protein shares high homology and typical structural features with other species SOs. Phylogenetic analysis indicates that NbSO cDNA clone encodes a tobacco SO isoform. Southern blot analysis suggests that NbSO is a single-copy gene in the N. benthamiana genome. The NbSO transcript levels were higher in aerial tissues and were up-regulated in N. benthamiana during sulfite stress. Reducing the SO expression levels through virus-induced gene silencing caused a substantial accumulation in sulfite content and less sulfate accumulation in N. benthamiana leaves when exposed to sulfite stress, and thus resulted in decreased tolerance to sulfite stress. Taken together, this study improves our understanding on the molecular and functional properties of plant SO and provides genetic evidence on the involvement of SO in sulfite detoxification in a sulfite-oxidizing manner in N. benthamiana plants.

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Acknowledgments

This work was financially supported by the National Natural Science Foundation of China (30971548) and The Key Project of Science and Technology of Henan Tobacco Company (HYKJ201010). The authors thank Dr Daowen Wang (Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, China) for generously providing the PEBV-based pCAPE vector system.

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Authors and Affiliations

  1. College of Life Science, Henan Agricultural University, Zhengzhou, 450002, People’s Republic of China

    Zongliang Xia, Jianyu Wu, Ke Wu & Hua Zhang

  2. Key Laboratory of Physiology, Ecology and Genetic Improvement of Food Crops in Henan Province, Zhengzhou, 450002, People’s Republic of China

    Zongliang Xia

  3. Henan Tobacco Company, Zhengzhou, 450008, People’s Republic of China

    Xinhong Su

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  1. Zongliang Xia
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  2. Xinhong Su
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  3. Jianyu Wu
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  4. Ke Wu
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  5. Hua Zhang
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Correspondence to Zongliang Xia.

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Zongliang Xia and Xinhong Su contributed equally to this work.

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11033_2011_993_MOESM1_ESM.pdf

Fig. 1 Sequence alignment and phylogenetic analysis of sulfite oxidase (SO) from N. benthamiana and other species (A) An alignment is shown for the deduced amino acid sequence of SO from Nicotiana benthamiana (NbSO), Lycopersicon esculentum (LeSO), Solanum tuberosum (StSO), Arabidopsis thaliana (AtSO), Oryza sativa (OsSO), and Zea Mays (ZmSO). The numbers on the left indicate the amino acid position. Identical residues in all these proteins are shown in a black background. Dashes indicated gaps introduced for optimal alignment. The putative molybdopterin-binding domain and dimerization domain are underlined with a solid line and a dashed line, respectively. The boxed regions mark the predicted signal peptide. (B) Phylogenetic tree based on SO protein sequences from bacterial, plants, and animals. The bootstrap values shown were calculated based on 500 replications. The tree was constructed using the neighbor-joining method. NbSO, Nicotiana benthamiana (HQ699883); LeSO, Lycopersicon esculentum (DQ853413); StSO, Solanum tuberosum (DQ284487); AtSO, Arabidopsis thaliana (At3g01910); BoSO, Brassica oleracea (AC183495); OsSO, Oryza sativa (Os08g0530400); ZmSO, Zea Mays (FJ436404); PtSO, Populus trichocarpa (XP_002300104); HcSO, Hibiscus cannabinus (ACU33027); DmSO, Drosophila melanogaster (NM_133103); CeSO, Caenorhabditis elegans (NM_001029564); GgSO, Gallus gallus (P07850); HsSO, Homo sapiens (NM_000456); MmSO, Mus musculus (NM_173733); RnSO, Rattus norvegicus (NM_031127); EcSO, Escherichia coli (NP_288430). Fig. 2 Southern blot analysis of NbSO The genomic DNA samples from N. benthamiana were digested separately with either EcoRI, XbaI or BamHI. The hybridization bands are indicated by arrows. DNA size markers (HindIII digest of lambda DNA) are shown on the right. The results shown are typical of three independent hybridizations. (PDF 91 kb)

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Xia, Z., Su, X., Wu, J. et al. Molecular cloning and functional characterization of a putative sulfite oxidase (SO) ortholog from Nicotiana benthamiana . Mol Biol Rep 39, 2429–2437 (2012). https://doi.org/10.1007/s11033-011-0993-x

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