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Ectopic expression of SOD and APX genes in Arabidopsis alters metabolic pools and genes related to secondary cell wall cellulose biosynthesis and improve salt tolerance

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Abstract

Hydrogen peroxide (H2O2) is known to accumulate in plants during abiotic stress conditions and also acts as a signalling molecule. In this study, Arabidopsis thaliana transgenics overexpressing cytosolic CuZn-superoxide dismutase (PaSOD) from poly-extremophile high-altitude Himalayan plant Potentilla atrosanguinea, cytosolic ascorbate peroxidase (RaAPX) from Rheum australe and dual transgenics overexpressing both the genes were developed and analyzed under salt stress. In comparison to wild-type (WT) or single transgenics, the performance of dual transgenics under salt stress was better with higher biomass accumulation and cellulose content. We identified genes involved in cell wall biosynthesis, including nine cellulose synthases (CesA), seven cellulose synthase-like proteins together with other wall-related genes. RNA-seq analysis and qPCR revealed differential regulation of genes (CesA 4, 7 and 8) and transcription factors (MYB46 and 83) involved in secondary cell wall cellulose biosynthesis, amongst which most of the cellulose biosynthesis gene showed upregulation in single (PaSOD line) and dual transgenics at 100 mM salt stress. A positive correlation between cellulose content and H2O2 accumulation was observed in these transgenic lines. Further, cellulose content was 1.6–2 folds significantly higher in PaSOD and dual transgenic lines, 1.4 fold higher in RaAPX lines as compared to WT plants under stress conditions. Additionally, transgenics overexpressing PaSOD and RaAPX also displayed higher amounts of phenolics as compared to WT. The novelty of present study is that H2O2 apart from its role in signalling, it also provides mechanical strength to plants and aid in plant biomass production during salt stress by transcriptional activation of cellulose biosynthesis pathway. This modulation of the cellulose biosynthetic machinery in plants has the potential to provide insight into plant growth, morphogenesis and to create plants with enhanced cellulose content for biofuel use.

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Acknowledgements

This research was supported by grants from the Council of Scientific and Industrial Research (CSIR), New Delhi, India under CSIR Network Projects BSC107 and BSC109.

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Authors and Affiliations

  1. Department of Biotechnology, CSIR-Institute of Himalayan Bioresource Technology, Palampur, Himachal Pradesh, India

    Amrina Shafi, Paramvir Singh Ahuja, Yelam Sreenivasulu, Sanjay Kumar & Anil Kumar Singh

  2. National Institute of Arthritis and Musculoskeletal and Skin Diseases, National Institute of Health, Bethesda, MD, USA

    Tejpal Gill

  3. Bioinformatics Centre, University of Kashmir, Srinagar, Jammu & Kashmir, India

    Insha Zahoor

  4. ICAR-Indian Institute of Agricultural Biotechnology, Ranchi, India

    Anil Kumar Singh

  5. Biotechnology Department, University of Kashmir, Srinagar, 190006, Jammu & Kashmir, India

    Amrina Shafi

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  1. Amrina Shafi
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  2. Tejpal Gill
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Contributions

AKS, AS and PSA conceived and designed the experiments. AS and TG performed the experiments. PSA, YS, SK and AKS analyzed the data. IZ did statistical analysis. AS and AKS wrote the manuscript.

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Correspondence to Amrina Shafi or Anil Kumar Singh.

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11033_2019_4648_MOESM1_ESM.pdf

Hierarchical tree graph of over-represented GO terms in down-regulated genes by singular enrichment analysis generated by agriGO. Boxes in the graph show GO terms labelled by their GO ID, term definition and statistical information. The significant terms (adjusted p  < 0.05) are marked with colour, while non-significant terms are shown as white boxes. The degree of colour saturation of a box correlates positively with the enrichment level of the term. Solid, dashed and dotted lines represent two, one and zero enriched terms at both ends connected by the line, respectively. The rank direction of the graph runs from top to bottom. (PDF 500 KB)

11033_2019_4648_MOESM2_ESM.pdf

UPLC Analysis of Soluble Phenolics in WT (a), S26 (b), APX20 (c) and 18O (d) lines of Arabidopsis thaliana. Phenolic profiles of transgenic (b, c, d) and wild type (a) lines were compared at 270 nm under control and stress conditions. Key to peak Peaks was confirmed by mass spectrometry. (PDF 267 KB)

Primer sequence and PCR conditions for genes used for semi-quantitative and real-time expression analysis. (DOCX 28 KB)

11033_2019_4648_MOESM4_ESM.xlsx

RNA seq FPKM values for the genes and transcription factors involved in secondary cell wall cellulose biosynthesis. (XLSX 22 KB)

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Shafi, A., Gill, T., Zahoor, I. et al. Ectopic expression of SOD and APX genes in Arabidopsis alters metabolic pools and genes related to secondary cell wall cellulose biosynthesis and improve salt tolerance. Mol Biol Rep 46, 1985–2002 (2019). https://doi.org/10.1007/s11033-019-04648-3

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