Extended Data Fig. 5: Validation of amino acid and nucleotide synthesis gene knockouts in breast cancer cells.
From: Nutrient requirements of organ-specific metastasis in breast cancer
a, Schematic of amino acid and nucleotide synthesis pathways, highlighting enzymes targeted to generate auxotrophs. Genes knocked out in breast cancer cells are shown in red; relevant metabolites synthesized are shown in blue. Asp: aspartate; DHO: dihydroorotate; Glu: glutamate; Gln: glutamine; IMP: inosine monophosphate; P5C: 1-pyrroline-5-carboxylic acid; PRPP: phosphoribosyl diphosphate; UMP: uridine monophosphate. Created in BioRender. Abbott, K. (2025) https://BioRender.com/lz9ywec. b, Western blot analysis of parental MDA-MB-231, EO771, and HCC1806 breast cancer cells for expression of the indicated proteins. c, Quantification of western blots in (b). Signal intensity of each protein was normalized to vinculin loading control. Data are mean ± SD; n = 3 biologically independent samples. Statistical analysis was performed using one-way ANOVA with Holm–Sidak multiple comparisons test (two-sided). d-i, Western blot analysis of non-targeting control (Ctrl) or knockout (KO) auxotroph clonal cells for expression of the indicated proteins in MDA-MB-231, HCC1806, or EO771 cells. PYCR KO represents PYCR1/2/3 triple KO. For gel source data, see Supplementary Fig. 1. Representative blots are shown from one of two independent experiments with similar results.
