Metabolite Extraction and Analysis Protocol

Metabolite extraction and analysis was performed as previously described^{10 (link), 29 (link)}. Briefly, cells were washed with PBS once and extracted with ice-cold extraction solvent (Acetonitrile/MeOH/ H₂O (30/50/20)), shaken for 5 min at 4 °C, transferred into eppendorf tubes and centrifuged for 5 min at 18k G. The supernatant was transferred in LC-MS glass vials and kept at −80 °C until measurement. Formate extraction and derivatization was performed as described in^{10 (link)} using a Methylchloroformate derivatization approach. Derivatized formate was analyzed using GC-MS (Agilent). Heavy labeled M + 2 formate was used as internal standard for quantification.
LC-MS analysis was performed as described previously^{10 (link)} using HILIC chromatography and a Q-Exactive mass spectrometer (Thermo Fisher Scientific). Raw data analysis was performed using TraceFinder (Thermo Fisher Scientific) software. Peak areas were normalized to cell volume in case of in vitro experiments, or to wet weight (mg) tissue.