Formaldehyde assimilation through coordination of the glyoxylate pathway and the tricarboxylic acid cycle in broad bean roots

Abstract

Formaldehyde (HCHO) assimilation in broad bean (Vicia faba L. cv. YD) roots was investigated using 13C-labeled HCHO followed by 13C-NMR analysis. Results revealed that H13CHO was first oxidized to H13COOH in the roots treated with 2 mM H13CHO in a time-dependent manner. Subsequently, a massive signal peak of [2, 4–13C]citrate (Cit) and a signal peak of [2, 3–13C]succinate (Su) were observed in accompany with an enhancement in the signal intensity of [3–13C]Cit. The data suggested that the glyoxylate pathway and the tricarboxylic acid (TCA) cycle functioned simultaneously in the subsequent assimilation of H13COOH. The yield of [2, 4–13C]Cit accounted for more than 80% of the total metabolites. The activity of isocitrate lyase (ICL), a key enzyme in the glyoxylate pathway, was stimulated by HCHO in a dosage-dependent manner. As a result, [2, 4–13C]Cit production was increased significantly in YD roots treated with high concentrations (4 and 6 mM) of H13CHO. Moreover, induction of the ICL activity by methanol resulted in a simultaneous elevation in the production of [2, 4–13C]Cit and [3–13C]Cit in methanol-pretreated roots under 2 mM H13CHO stress. Pretreatment of roots with cyclosporin A, which hinders the transport of 13C-enriched compounds into mitochondria, caused a notable decline in the signal peak and yield of [2, 4–13C]Cit and consequently induced a notable accumulation of [2, 3–13C]Su and an increase in the HCO3− production (generated from H13COOH oxidation) in H13CHO-treated roots. These results suggested that the glyoxylate pathway and the TCA cycle function coordinately in HCHO assimilation in broad bean roots.