Figure 3. PLCγ1 suppression decreases mitochondrial respiration and enhances cancer cell glycolytic capacity.
(a) ER Ca2+ release rate (n = 9/11), intraluminal content (n = 9), cytosolic and mitochondrial Ca2+ response (n = 10) in cells transduced with either empty vector (shControl) or doxycycline-inducible shRNA against PLCγ1.
(b) Oxygen consumption rate (OCR) in A549 cells transduced as in (a), incubated with doxycycline for 48h and moved to normoxia or hypoxia for 48h; n = 8.
(c) Extracellular acidification rate (ECAR) in A549 cells transduced and treated as in (b); n normoxia/hypoxia = 9/8.
(d) ECAR in A549 cells transfected with either pcDNA3.1 empty vector or pcDNA3.1-PLCγ1 and incubated for 48h in normoxia or hypoxia; n = 7.
(e) ECAR in MEFs of the indicated genotype transfected with either pcDNA3.1-HA-LIC empty vector or pcDNA3.1-HA-LIC-PLCγ1 and incubated for 48h in hypoxia; n = 6.
(f) β-oxidation of cells transduced with either an empty vector (shControl) or doxycycline-inducible shRNAs against PLCγ1, transfected with pcDNA3.1 empty vector or pcDNA3.1-rPLCγ1 (shRNA-resistant PLCγ1) and incubated for 48h in hypoxia; n = 4.
(g) β-oxidation of cells transfected with either pcDNA3.1 empty vector or pcDNA3.1-PLCγ1 and incubated for 48h in normoxia or hypoxia; n = 4.
(h) Triacylglycerols measured by ultra performance liquid chromatography - tandem mass spectrometry of cells treated as in (b); n = 4/group.
(i) Lipid droplet quantification by flow cytometry (relative to normoxia) of cells transduced as in (b) and stained with LipidTOX (far red); n = 3
(j) Representative confocal fluorescence microscopy images of lipid droplet staining with LipidTOX (far red) in cells treated as in (b). Hoechst: nuclei.
(k) Representative quantification of LD area/cell from (j); n = 13 biologically independent replicates (50 cells/replicate).
(l) Lipid droplet quantification (relative to normoxia) by flow cytometry (left) and representative flow cytometry panel (right) of cells treated as indicated and stained with LipidTOX; n = 3.
Graphical data are mean ± SD. Statistical analyses were done using two-tailed unpaired Student’s t test or one-way ANOVA; n, number of biologically independent samples. **** p < 0.0001. Statistical source data are provided in Source Data Fig. 3.