Cell-selective labeling using amino acid precursors for proteomic studies of multicellular environments

• Published in: Nature methods Vol. 10; no. 8; pp. 768 - 773
• Main Authors: Gauthier, Nicholas P, Soufi, Boumediene, Walkowicz, William E, Pedicord, Virginia A, Mavrakis, Konstantinos J, Macek, Boris, Gin, David Y, Sander, Chris, Miller, Martin L
• Format: Journal Article
• Language: English
• Published: New York Nature Publishing Group US 01.08.2013; Nature Publishing Group
• Subjects: 631/1647/2067; 631/45/475; 631/80/86; 692/308/1892; Affinity labeling; Amino acids; Animals; Base Sequence; Bioinformatics; Biological Microscopy; Biological Techniques; Biomarkers; Biomedical Engineering/Biotechnology; Biosynthesis; Cell Line; Cellular biology; Coculture Techniques - methods; Enzymes; Humans; Isotope Labeling - methods; Life Sciences; Lysine - metabolism; Mass spectrometry; Methods; Mice; Molecular Sequence Data; Oligonucleotide Array Sequence Analysis; Organisms, Genetically Modified; Proteins; Proteome - biosynthesis; Proteome - genetics; Proteomics; Proteomics - methods; RNA, Messenger - chemistry; RNA, Messenger - genetics; Sequence Analysis, DNA; Tandem Mass Spectrometry; Vertebrates; United States; Germany
• ISSN: 1548-7091; 1548-7105
• DOI: 10.1038/nmeth.2529

A cellular engineering approach coupled with mass spectrometry allows the cell-of-origin of intra- and extracellular proteins to be determined from co-cultured cells. We report a technique to selectively and continuously label the proteomes of individual cell types in coculture, named cell type–specific labeling using amino acid precursors (CTAP). Through transgenic expression of exogenous amino acid biosynthesis enzymes, vertebrate cells overcome their dependence on supplemented essential amino acids and can be selectively labeled through metabolic incorporation of amino acids produced from heavy isotope–labeled precursors. When testing CTAP in several human and mouse cell lines, we could differentially label the proteomes of distinct cell populations in coculture and determine the relative expression of proteins by quantitative mass spectrometry. In addition, using CTAP we identified the cell of origin of extracellular proteins secreted from cells in coculture. We believe that this method, which allows linking of proteins to their cell source, will be useful in studies of cell-cell communication and potentially for discovery of biomarkers.