Metabolomic analysis ofArabidopsisreveals hemiterpenoid glycosides as products of a nitrate ion-regulated, carbon flux overflow

Abstract

An understanding of the balance between carbon and nitrogen assimilation in plants is key to future bioengineering for a range of applications. Metabolomic analysis of the model plant, Arabidopsis thaliana, using combined NMR-MS revealed the presence of two hemiterpenoid glycosides that accumulated in leaf tissue, to ∼1% dry weight under repeated nitrate-deficient conditions. The formation of these isoprenoids was correlated with leaf nitrate concentrations that could also be assayed in the metabolomic data using a unique flavonoid-nitrate mass spectral adduct. Analysis of leaf and root tissue from plants grown in hydroponics with a variety of root stressors identified the conditions under which the isoprenoid pathway in leaves was diverted to the hemiterpenoids. These compounds were strongly induced by root wounding or oxidative stress and weakly induced by potassium deficiency. Other stresses such as cold, saline, and osmotic stress did not induce the compounds. Replacement of nitrate with ammonia failed to suppress the formation of the hemiterpenoids, indicating that nitrate sensing was a key factor. Feeding of intermediates was used to study aspects of 2-C-methyl-D-erythritol-4-phosphate pathway regulation leading to hemiterpenoid formation. The formation of the hemiterpenoids in leaves was strongly correlated with the induction of the phenylpropanoids scopolin and coniferin in roots of the same plants. These shunts of photosynthetic carbon flow are discussed in terms of overflow mechanisms that have some parallels with isoprene production in tree species.F or centuries mankind has relied on carbon, prehistorically fixed from the atmosphere by plants and algae, as a source of fuel and other materials. This is a dwindling resource and, to develop renewable sources of chemicals, knowledge of the biochemical pathways that link photosynthesis to the accumulation of useful materials will be paramount. Terpenoids (isoprenoids) are examples of carbon-rich products that occur prolifically across the plant kingdom. They are not only end products of chloroplast assimilation of photosynthetic carbon but also, by virtue of the phytyl side chains of chlorophyll, key elements of the machinery of photosynthesis. Some terpenoids are ubiquitous and are classed as primary metabolites (sterols, carotenoids, and phytyl side chains of chlorophyll); others are hormones (gibberellins, abscisic acid, brassinosteroids, and strigolactones). However, the majority of terpenoids, especially those that can accumulate to high levels in plants, are classed as secondary metabolites, although many have roles in chemical ecology. Manipulation of flux through the primary terpenoid pathways and the accumulation of terpenoids via the secondary pathways both have potential for exploitation. Arabidopsis, the model plant, was, until recently, considered to be poorly endowed with terpenoids. For example, this species does not appear to contain oxidized sesqui-and diterpenes, compounds that are widespread in plants (1). There are some 30 terpene synt...

Deoxyxylulose 5-Phosphate Synthase Controls Flux through the Methylerythritol 4-Phosphate Pathway in Arabidopsis

“…MEcDP is normally converted to HMBDP en route to IDP and DMADP production by HMBDP reductase (HDR) in the stroma. However, root wounding, oxidative stress, and nitrate deficiency in Arabidopsis leaves can trigger diversion of HMBDP into glycosylated hemiterpenoids (Ward et al, 2011).…”

Cell Metabolomics

“…Cell metabolomics is an emerging field that addresses fundamental biological questions and allows observation of metabolic phenomena in cells. Thus, metabolomics has been used for optimizing cell cultures for antibody production (Wu et al, 2012), testing drugs (Sabidó et al, 2012), cell transfections, determinant of apoptosis, and comparing lung cancer cell phenotypes, identifying novel underlying metabolic pathways (Ward et al, 2011).…”

The diversion of 2‐C‐methyl‐d‐erythritol‐2,4‐cyclodiphosphate from the 2‐C‐methyl‐d‐erythritol 4‐phosphate pathway to hemiterpene glycosides mediates stress responses in Arabidopsis thaliana

“…This sequence can be interrupted by certain types of stress which cause diversion of some pathway intermediates to other metabolic fates. For instance, HMBDP can be converted to hemiterpene glycosides in Arabidopsis (Figure 1) as a response to root wounding, NO 3 À deficiency, or cold stress (Ward et al, 2011). However, the details of such conversions, such as the enzymes which catalyze these processes or the subcellular compartments where such transformations take place, are currently unknown.…”