Abstract
Carotenoids and chlorophylls are photosynthetic pigments synthesized in plastids from metabolic precursors provided by the methylerythritol 4-phosphate (MEP) pathway. The first two steps in the MEP pathway are catalyzed by the deoxyxylulose 5-phosphate synthase (DXS) and reductoisomerase (DXR) enzymes. While DXS has been recently shown to be the main flux-controlling step of the MEP pathway, both DXS and DXR enzymes have been proven to be able to promote an increase in MEP-derived products when overproduced in diverse plant systems. Carrot (Daucus carota) produces photosynthetic pigments (carotenoids and chlorophylls) in leaves and in light-exposed roots, whereas only carotenoids (mainly α- and β-carotene) accumulate in the storage root in darkness. To evaluate whether DXS and DXR activities influence the production of carotenoids and chlorophylls in carrot leaves and roots, the corresponding Arabidopsis thaliana genes were constitutively expressed in transgenic carrot plants. Our results suggest that DXS is limiting for the production of both carotenoids and chlorophylls in roots and leaves, whereas the regulatory role of DXR appeared to be minor. Interestingly, increased levels of DXS (but not of DXR) resulted in higher transcript abundance of endogenous carrot genes encoding phytoene synthase, the main rate-determining enzyme of the carotenoid pathway. These results support a central role for DXS on modulating the production of MEP-derived precursors to synthesize carotenoids and chlorophylls in carrot, confirming the pivotal relevance of this enzyme to engineer healthier, carotenoid-enriched products.
Highlights
Many isoprenoids are present in plants and some of them act as primary metabolites with roles in respiration, photosynthesis, and regulation of growth and development
The first step in the methylerythritol 4-phosphate (MEP) pathway, which is catalyzed by the deoxyxylulose 5-phosphate (DXP) synthase (DXS) enzyme, is the formation of DXP from pyruvate and glyceraldehyde 3-phosphate
In order to determine the significance of the supply of MEPderived metabolic precursors for the synthesis of carotenoids in carrot, we generated transgenic plants expressing GFP-tagged versions of the Arabidopsis deoxyxylulose 5-phosphate synthase (DXS)/DXS1/CLA1 (At4g15560) or DXP reductoisomerase (DXR) (At5g62790) genes under the control of the constitutive 35S promoter (Supplementary Figures S1 and S2)
Summary
Many isoprenoids are present in plants and some of them act as primary metabolites with roles in respiration, photosynthesis, and regulation of growth and development. Five more enzymes convert MEP (the first intermediate that is specific of this pathway) into IPP and DMAPP (Figure 1) (RodriguezConcepcion and Boronat, 2002). Different studies have shown that DXS has a major role in the control of the MEP pathway flux (Rodriguez-Concepcion and Boronat, 2015). Overexpression of DXS-encoding genes in different plants typically results in increased levels of plastidial isoprenoids such as carotenoids and chlorophylls (Estevez et al, 2001; Enfissi et al, 2005; CarreteroPaulet et al, 2006; Morris et al, 2006; Munoz-Bertomeu et al, 2006; Zhang et al, 2009; Henriquez et al, 2016). Overexpression of DXR-encoding genes led to increased levels of MEPderived isoprenoids in many cases (Mahmoud and Croteau, 2001; Carretero-Paulet et al, 2006; Hasunuma et al, 2008; Yang et al, 2012; Chang et al, 2014; Zhang et al, 2015) but had no effect in others (Mendoza-Poudereux et al, 2014)
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