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Characterization and functional analysis of eugenol O-methyltransferase gene reveal metabolite shifts, chemotype specific differential expression and developmental regulation in Ocimum tenuiflorum L.

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Abstract

Eugenol-O-methyltransferase (EOMT) catalyzes the conversion of eugenol to methyleugenol in one of the final steps of phenylpropanoid pathway. There are no comprehensive reports on comparative EOMT gene expression and developmental stage specific accumulation of phenylpropenes in Ocimum tenuiflorum. Seven chemotypes, rich in eugenol and methyleugenol, were selected by assessment of volatile metabolites through multivariate data analysis. Isoeugenol accumulated in higher levels during juvenile stage (36.86 ng g−1), but reduced sharply during preflowering (8.04 ng g−1), flowering (2.29 ng g−1) and postflowering stages (0.17 ng g−1), whereas methyleugenol content gradually increased from juvenile (12.25 ng g−1) up to preflowering (16.35 ng g−1) and then decreased at flowering (7.13 ng g−1) and post flowering (5.95 ng g−1) from fresh tissue. Extreme variations of free intracellular and alkali hydrolysable cell wall released phenylpropanoid compounds were observed at different developmental stages. Analyses of EOMT genomic and cDNA sequences revealed a 843 bp open reading frame and the presence of a 90 bp intron. The translated proteins had eight catalytic domains, the major two being dimerisation superfamily and methyltransferase_2 superfamily. A validated 3D structure of EOMT protein was also determined. The chemotype Ot7 had a reduced reading frame that lacked both dimerisation domains and one of the two protein-kinase-phosphorylation sites; this was also reflected in reduced accumulation of methyleugenol compared to other chemotypes. EOMT transcripts showed enhanced expression in juvenile stage that increased further during preflowering but decreased at flowering and further at postflowering. The expression patterns may possibly be compared and correlated to the amounts of eugenol/isoeugenol and methyleugenol in different developmental stages of all chemotypes.

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Acknowledgments

This work was supported by Departmental Grants of BIT Mesra. The authors are thankful to BTISNET SubDIC (BT/BI/04/065/04) for providing facilities for bioinformatics analyses and the Government of Jharkhand, Department of Agriculture (5/B.K.V/Misc/12/2001) for providing infrastructure development fund. Fellowships were provided to IKR by BIT-Mesra and IH by CSIR [9/554 (13) 2007-EMR-I].

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  1. Inamul Haque

    Present address: Department of Botany, Derozio Memorial College, Kolkata, 700136, India

  2. Amit Rai

    Present address: Department of Biological Science, National University of Singapore, Singapore, 117543, Singapore

Authors and Affiliations

  1. Department of Biotechnology, Birla Institute of Technology, Mesra, Ranchi, 835215, India

    Indu Kumari Renu, Inamul Haque, Manish Kumar, Raju Poddar, Rajib Bandopadhyay, Amit Rai & Kunal Mukhopadhyay

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  1. Indu Kumari Renu
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Correspondence to Kunal Mukhopadhyay.

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Indu Kumari Renu, Inamul Haque contributed equally.

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Renu, I.K., Haque, I., Kumar, M. et al. Characterization and functional analysis of eugenol O-methyltransferase gene reveal metabolite shifts, chemotype specific differential expression and developmental regulation in Ocimum tenuiflorum L.. Mol Biol Rep 41, 1857–1870 (2014). https://doi.org/10.1007/s11033-014-3035-7

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