Extending the Range of SLIM-Labeling Applications: From Human Cell Lines in Culture to Caenorhabditis elegans Whole-Organism Labeling.

The simple light isotope metabolic-labeling technique relies on the in vivo biosynthesis of amino acids from U-[ 12 C]-labeled molecules provided as the sole carbon source. The incorporation of the resulting U-[ 12 C]-amino acids into proteins presents several key advantages for mass-spectrometry-based proteomics analysis, as it results in more intense monoisotopic ions, with a better signal-to-noise ratio in bottom-up analysis. In our initial studies, we developed the simple light isotope metabolic (SLIM)-labeling strategy using prototrophic eukaryotic microorganisms, the yeasts Candida albicans and Saccharomyces cerevisiae , as well as strains with genetic markers that lead to amino-acid auxotrophy. To extend the range of SLIM-labeling applications, we evaluated (i) the incorporation of U-[ 12 C]-glucose into proteins of human cells grown in a complex RPMI-based medium containing the labeled molecule, considering that human cell lines require a large number of essential amino-acids to support their growth, and (ii) an indirect labeling strategy in which the nematode Caenorhabditis elegans grown on plates was fed U-[ 12 C]-labeled bacteria ( Escherichia coli ) and the worm proteome analyzed for 12 C incorporation into proteins. In both cases, we were able to demonstrate efficient incorporation of 12 C into the newly synthesized proteins, opening the way for original approaches in quantitative proteomics.