Extended Data Fig. 6: Validation of auxotroph cells by assessing proliferation with or without rescue metabolites.
From: Nutrient requirements of organ-specific metastasis in breast cancer
a-f, Percent confluence of control (Ctrl) or knockout (KO) clonal cells cultured in medium with or without the relevant rescue metabolites. ASNS KO (a) and PYCR KO (d) were cultured in DMEM; ASS1 KO (b) and PHGDH KO (c) in RPMI lacking arginine or serine, respectively; DHODH KO (e) and GART KO (f) in RPMI. Rescue metabolite concentrations: 1.15 mM arginine (Arg), 379 µM asparagine (Asn), 1 mM citrulline (Cit), 100 µM hypoxanthine (Hx), 174 µM proline (Pro), 286 µM serine (Ser), 100 µM uridine (Urd). Data are mean ± SD; n = 3 biologically independent samples. g, Percent confluence of EO771 cells in standard RPMI (11 mM glucose), RPMI containing 25 mM glucose (DMEM-equivalent), or standard RPMI refreshed every 48 h. Data are mean ± SD; n = 3 biologically independent samples. h-k, Percent confluence of HCC1806 Ctrl or auxotroph KO cells (ASNS KO, ASS1 KO, PHGDH KO, PYCR KO) cultured in DMEM lacking the indicated metabolites. h, ± Asn. i, Without Arg, ± Cit. j, ± Ser. k, ± Pro. Rescue metabolite concentrations: 398 µM Arg, 379 µM Asn, 1 mM Cit, 174 µM Pro, 400 µM Ser. Data are mean ± SD; n = 3 biologically independent samples. Asn, asparagine; Arg, arginine; Cit, citrulline; Ser, serine; Pro, proline; Urd, uridine; Hx, hypoxanthine; PYCR KO, PYCR1/2/3 triple KO. Representative plots are shown from one of two independent experiments with similar results.
