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Redirection of metabolite biosynthesis from hydroxybenzoates to volatile terpenoids in green hairy roots of Daucus carota

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Abstract

Main conclusion

A metabolic shift in green hairy root cultures of carrot from phenylpropanoid/benzenoid biosynthesis toward volatile isoprenoids was observed when compared with the metabolite profile of normal hairy root cultures.

Hairy roots cultures of Daucus carota turned green under continuous illumination, while the content of the major phenolic compound p-hydroxybenzoic acid (p-HBA) was reduced to half as compared to normal hairy roots cultured in darkness. p-Hydroxybenzaldehyde dehydrogenase (HBD) activity was suppressed in the green hairy roots. However, comparative volatile analysis of 14-day-old green hairy roots revealed higher monoterpene and sesquiterpene contents than found in normal hairy roots. Methyl salicylate content was higher in normal hairy roots than in green ones. Application of clomazone, an inhibitor of 1-deoxy-d-xylulose 5-phosphate synthase (DXS), reduced the amount of total monoterpenes and sesquiterpenes in green hairy roots compared to normal hairy roots. However, methyl salicylate content was enhanced in both green and normal hairy roots treated with clomazone as compared to their respective controls. Because methyl-erythritol 4-phosphate (MEP) and phenylpropanoid pathways, respectively, contribute to the formation of monoterpenes and phenolic acids biosynthesis, the activities of enzymes regulating those pathways were measured in terms of their in vitro activities, in both green and normal hairy root cultures. These key enzymes were 1-deoxy-d-xylulose 5-phosphate reductoisomerase (DXR), an early regulatory enzyme of the MEP pathway, pyruvate kinase (PK), an enzyme of primary metabolism related to the MEP pathway, shikimate dehydrogenase (SKDH) which is involved in biosynthesis of aromatic amino acids, and phenylalanine ammonia-lyase (PAL) that catalyzes the first step of phenylpropanoid biosynthesis. Activities of DXR and PK were higher in green hairy roots as compared to normal ones, whereas the opposite trend was observed for SKDH and PAL activities. Gene expression analysis of DXR and PAL showed trends similar to those for the respective enzyme activities. Based on these observations, we suggest a possible redirection of metabolites from the primary metabolism toward isoprenoid biosynthesis, limiting the phenolic biosynthetic pathway in green hairy roots grown under continuous light.

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Abbreviations

DXR:

1-Deoxy-d-xylulose 5-phosphate reductoisomerase

DXS:

1-Deoxy-d-xylulose 5-phosphate synthase

EPSPS:

5-Enolpyruvyl-shikimate-3-phosphate synthase

HBD:

p-Hydroxybenzaldehyde dehydrogenase

MEP:

Methyl-erythritol 4-phosphate

MVA:

Mevalonic acid

PAL:

Phenylalanine ammonia-lyase

p-HBA:

p-Hydroxybenzoic acid

PK:

Pyruvate kinase

SKDH:

Shikimate dehydrogenase

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Acknowledgments

This work was supported by a research grant [No. 38(1201)/08/EMR-II to A. Mitra] from the Council of Scientific and Industrial Research (CSIR), India. Chiranjit Mukherjee was supported from this grant in the form of a senior research fellow. Facilities created from a research grant (4-25/2013/TS-I) of Ministry of Human Resource Development (MHRD) India were utilized in this work. Finally, we thank Ishita Paul, a member of our research group, for checking grammatical errors.

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  1. Natural Product Biotechnology Group, Agricultural and Food Engineering Department, Indian Institute of Technology Kharagpur, Kharagpur, 721 302, India

    Chiranjit Mukherjee, Tanmoy Samanta & Adinpunya Mitra

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  1. Chiranjit Mukherjee
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Correspondence to Adinpunya Mitra.

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Mukherjee, C., Samanta, T. & Mitra, A. Redirection of metabolite biosynthesis from hydroxybenzoates to volatile terpenoids in green hairy roots of Daucus carota . Planta 243, 305–320 (2016). https://doi.org/10.1007/s00425-015-2403-4

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