Extended Data Fig. 5: Asparagine Rescues the Inhibition of Respiration.
A. Relative confluence of clone N treated with 1 nM oligomycin in the presence or absence of media containing a 100 µM cocktail of all NEAAs after 5 days (n = 3). B. Relative confluence of clone N treated with 1 nM oligomycin in the presence or absence of individual NEAAs after 5 days (n = 3). C,D. Dose dependent rescue of doubling time (C) and confluence after 5 days of culture (D) by exogenous asparagine (Asn) following treatment with 1 nM oligomycin. E. Relative confluence of clone N treated with 1 nM oligomycin in the presence or absence of 100 µM or 20 mM aspartate (n = 3). F. Relative confluence of clonal line N treated with 1 nM oligomycin, 25 µM phenformin, or 25 nM IACS-10759 after 5 days + /- Asn (n = 3). G. Immunoblot analysis of siRNA targeting Asns or non-targeted (NT) control in clonal line N or V. H,I. Representative images of vehicle vs 1 nM oligomycin treated co-cultures of labeled clone N with unlabeled clone N (H) or insensitive clone V (I) transfected with the indicated siRNA with or without the addition of exogenous Asn. Error bars are mean ± SD, Scale Bars = 400 µm, * P ≤ 0.05; ** P ≤ 0.01; *** P ≤ 0.001; **** P ≤ 0.0001 by two-tailed student’s t test (A,F), or one-way Anova with Tukey post hoc (B,E).
