Figure 1.

Characterization, genetic modification and proteome-labelling strategy of MSCs. (A) FACS analysis of MSCs for cell surface markers. (B) MSCs under a phase-contrast microscope (Left panel) and after differentiation in culture into adipocytes (Middle panel, oil red staining) and osteocytes (Right panel, alizarin red staining). Shown are representatives of three independent experiments. (C) MSCs were infected with lentiviral vector MetRS^L274G-mCherry, and the transduced cells were FACS-selected by mCherry expression. (D) Expression of MetRS^L274G allows loading of the methionine surrogate azidonorleucine (ANL) onto methionine tRNA, and subsequently ANL incorporation in newly-synthesized proteins (Upper panel). The ANL-tagged proteins are clicked to alkyne-containing dye or resin in the presence of copper, and detected by FUNCAT/BONCAT or enriched for LC-MS (Lower panel).