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Nitrogen sources are all converted into ammonium/ia as a first step of assimilation. It is reasonable to expect that molecular components involved in the transport of ammonium/ia across biological membranes connect with the regulation of both nitrogen and central metabolism. We applied both genetic (i.e., Δamt mutation) and environmental treatments to a target biological system, the cyanobacterium Anabaena sp PCC 7120. The aim was to both perturb nitrogen metabolism and induce multiple inner nitrogen states, respectively, followed by targeted quantification of key proteins, metabolites and enzyme activities. The absence of AMT transporters triggered a substantial whole-system response, affecting enzyme activities and quantity of proteins and metabolites, spanning nitrogen and carbon metabolisms. Moreover, the Δamt strain displayed a molecular fingerprint indicating nitrogen deficiency even under nitrogen replete conditions. Contrasting with such dynamic adaptations was the striking near-complete lack of an externally measurable altered phenotype. We conclude that this species evolved a highly robust and adaptable molecular network to maintain homeostasis, resulting in substantial internal but minimal external perturbations. This analysis provides evidence for a potential role of AMT transporters in the regulatory/signalling network of nitrogen metabolism and the existence of a novel fourth regulatory mechanism controlling glutamine synthetase activity., (© 2021 The Authors. Plant, Cell & Environment published by John Wiley & Sons Ltd.)

The universality of low molecular weight metabolites allows rapid and straightforward investigation of the biochemistry of genetically uncharacterised species. Thus ex vivo metabolic profiling in combination with multivariate data analysis (metabonomics) offers great potential in comparative biology. Here we present the first use of high resolution nuclear magnetic resonance (NMR) spectroscopy to distinguish closely related animal species via their metabolic phenotype (metabotype). We have profiled the three Eisenia (Oligochaeta, Lumbricidae) species Eisenia fetida, Eisenia andrei and Eisenia veneta using tissue extracts and coelomic fluid analysis. The low molecular weight biochemical profiles of tissue extracts were highly conserved for all three species, with E. fetida and E. andrei being more similar to each other than to E. veneta. However the metabolic profiles of the coelomic fluid of the different species were highly distinctive - the NMR spectra allowed unequivocal identification of species. Multivariate statistics were also used to quantify these spectral differences and to enable simplified graphical visualisation of species similarity. These results show that two morphologically undistinguishable species (E. fetida and E. andrei) differ markedly in their biochemical profiles despite apparently occupying the same ecological niche, and indicate that metabolic phenotype profiling can be used as a powerful functional genomics tool.

The endogenous metabolites of the coelomic fluid of the earthworm Eisenia veneta were characterised using high-resolution one-dimensional and two-dimensional 1H nuclear magnetic resonance spectroscopy. Signals from common organic acids, such as acetate, fumarate, malonate, malate, formate, and succinate, were identified together with adenosine and nicotinamide mononucleotide. The potential use of this information as a baseline dataset for future toxicological or physiological studies was demonstrated by a metabonomic analysis: a series of earthworms were dosed with the model compound 3-fluoro-4-nitrophenol, and toxic effects followed by multivariate analysis of the spectral data of the coelomic fluid. Relative concentrations of acetate and malonate were decreased in the dosed worms compared to the controls.