Fig. 3: Mtb infected GID/CTLH knockout macrophages have altered metabolism and are strongly glycolytic.

A Flux analyses of scramble or GID8^−/−, MAEA^−/− and WDR26^−/− BMDM^hs infected with Mtb Erdman strain at MOI 1 24 hours post infection. Oxygen consumption rates (OCR) were measured using the Cell Mito Stress Test Kit (Agilent). Oligo, oligomycin; FCCP, fluoro-carbonyl cyanide phenylhydrazone; Rot/A, rotenone and antimycin A. B Comparison of basal respiration and spare respiratory capacity (SRC) from (A). SRC was calculated by subtracting the normalized maximal OCR from basal OCR; n = 3 biological replicates (two technical replicates per repeat). ****P < 0.0001, one-way ANOVA alongside Dunnett’s multiple comparison test. Data are presented as mean values ± SD. C Extracellular acidification rates (ECARs) of scramble or GID8^−/−, MAEA^−/− and WDR26^−/− BMDM^hs infected with Mtb as in (A). ECARs were measured using the Agilent Seahorse Glycolysis Stress Test kit. 2DG, 2-Deoxy-D-glucose. D Comparison of basal glycolysis and spare glycolytic capacity (SGC) in the indicated BMDM^h knockouts. SGC was calculated as SRC above. n = 3 biological replicates (two technical replicates per repeat). **P < 0.01; ****P < 0.0001, one-way ANOVA alongside Dunnett’s multiple comparison test. Data are presented as mean values ± SD. E Western blot analysis of AMPK phosphorylation in scramble or GID8^−/−, MAEA^−/−, and WDR26^−/− BMDM^hs uninfected or infected with Mtb at MOI 4. F Quantification of relative expression of p-AMPK in (E) and in Fig. S8A, B normalized to β-actin; n = 3 biological replicates. *P < 0.05; **P < 0.01, two-way ANOVA alongside Dunnett’s multiple comparison test. Data are presented as mean values ± SD. G p-AMPK/AMPK ratios of relative expressions in (E) and in Fig. S8A, B as normalized to β-actin; n = 3 biological replicates. ***P < 0.001; ****P < 0.0001, two-way ANOVA alongside Dunnett’s multiple comparison test. Data are presented as mean values ± SD. Source data are provided as a Source Data file.