Extended Data Fig. 2: Glutamine de novo synthesis can maintain collagen synthesis and proliferation when glutamine is limiting.
From: Fibroblast pyruvate carboxylase is required for collagen production in the tumour microenvironment
(A) Western Blot of PSCs cultured in 20% Gln in the presence of TGFβ and treated with dm-αKG and MSO. (B) Collagen abundance in ECM derived from confluent PSCs cultured in 100% or 10% Gln in the presence of TGFβ and treated with dm-αKG, dm-Glu and MSO. n = 3 biologically independent samples. (C) Relative number of NIH-3T3 cells expressing Ctrl or Glul sgRNA, cultured in 10% Gln and treated with TGFβ alone and dm-αKG or dm-Glu. n = 2 (Glul sg6 dm-Glu), n = 3 (all others) biologically independent samples. (D,E) Growth curves of NIH-3T3 cells (D) or PSCs (E) expressing Ctrl or Glul sgRNA, cultured in 100% or 10% Gln/20% Gln. n = 3 biologically independent samples. (F) Western Blot of PSCs expressing Ctrl or Glul sgRNA, cultured in 20% Gln for 48 h. (G) Collagen abundance in ECM derived from confluent PSCs expressing Ctrl or Glul sgRNA, cultured in 20% Gln. n = 3 biologically independent samples. Mean+SD (B,C,G) or mean±SD (D,E) are shown. One-way ANOVA with Holm-Sidak correction (B), one-way ANOVA (C,G), two-sided unpaired t-test (C: Glul sg6 dm-Glu), two-way ANOVA (D,E). Western blots (A,F) are representative of three independent experiments. All other experiments were performed at least twice.
