Profiling the Metabolism of Human Cells by Deep ¹³C Labeling

Abstract

The metabolism of most human cell types remains largely unexplored, in part due to lack of methods for measuring metabolic activities in an unbiased, hypothesis-free manner. Here we introduce the deep labeling method for profiling metabolic activities, based on a custom ¹³C medium in combination with high-resolution mass spectrometry. A proof-of-principle study on human cancer cells revealed hundreds of endogenous metabolites and identified a variety of metabolic activities, but also lack of central pathways synthesizing cysteine, carnitine and creatine. Protein and nucleic acids were almost exclusively de novo synthesized, while lipids were partly derived from serum fat. Branched-chain keto acids (BCKA) were formed and metabolized to a short-chain acylcarnitines, but did not enter the TCA cycle. Remarkably, BCKA could substitute for essential amino acids to support growth of these cells. We anticipate that deep labeling can be used to catalogue metabolic phenotypes across a range of cell types and conditions.