Metabotyping of the C. elegans sir-2.1 Mutant Using in Vivo Labeling and 13C‑Heteronuclear Multidimensional NMR Metabolomics

The roles of sir-2.1 in C. elegans lifespan extension have been subjects of recent public and academic debates. We applied an efficient workflow for in vivo 13C-labeling of C. elegans and 13C-heteronuclear NMR metabolomics to characterizing the metabolic phenotypes of the sir-2.1 mutant. Our method delivered sensitivity 2 orders of magnitude higher than that of the unlabeled approach, enabling 2D and 3D NMR experiments. Multivariate analysis of the NMR data showed distinct metabolic profiles of the mutant, represented by increases in glycolysis, nitrogen catabolism, and initial lipolysis. The metabolomic analysis defined the sir-2.1 mutant metabotype as the decoupling between enhanced catabolic pathways and ATP generation. We also suggest the relationship between the metabotypes, especially the branched chain amino acids, and the roles of sir-2.1 in the worm lifespan. Our results should contribute to solidifying the roles of sir-2.1, and the described workflow can be applied to studying many other proteins in metabolic perspectives.