Extended Data Fig. 1: Expression analysis of UCP2 and UCP2 homologs in PDAC cells.
a, Transcript abundance of UCP2 in PDAC primary tumour samples and normal pancreatic tissues derived from gene expression profiling (GEP) of different publicly available datasets. b–d, The relative expression of UCP2 in control and UCP2shRNA-transfected PDAC cells. UCP2 transcript levels were quantified by qPCR (b and d) and UCP2 protein levels were assayed with a UCP2-specific antiserum (c). An anti-COX4 antibody was used for normalization. e, f, UCP2 silencing does not alter the expression levels of other UCP homologs. The relative expression of all UCP genes was quantified by qPCR. The ΔCT of the indicated UCP homolog gene (e) and that of UCP2 (f) of the CtrshRNA-transfected cells were used as internal calibrators. g, ATP/ADP and Pi/Asp exchanges activities catalysed by mitochondrial extracts of control, UCP2-silenced and UCP2-silenced rescued PDAC cells. Mitochondria were solubilized with detergent and reconstituted into liposomes containing Pi or ATP. The exchange reaction was started by adding [14C]aspartate or [14C]ADP to proteoliposomes and stopped after 30 minutes with inhibitors. Values represent means ± SD of three independent experiments. h, Expression of murine Ucp2 (mUcp2) gene in Patu8988T cells. The relative expression of human UCP2 and murine Ucp2 were determined by Sybr green qPCR. The ΔCT of human UCP2 of the CtrshRNA-transfected cells carrying the empty expression vector (EV) was used as internal calibrator. The protein expression levels of panel h are shown in panel c. i, Effect of glutamine on UCP2 protein expression levels. The total cell extracts were assayed in the presence or absence (overnight starvation) of glutamine. For all transcript analyses, reverse transcribed cDNA from three biological replicates was used and three technical replicates were analysed for each biological replicate, values represent means ± SD (b, d–f, h). COX4 normalization was carried out on the same SDS-PAGE used to assay UCP2 expression levels, similar results were obtained in three biologically independent experiments (c, i). Statistical significance was calculated by unpaired two-tailed Mann–Whitney U-test (a). rUCP2, recombinant human UCP2. Patu8988TNI, cells transfected with non-inducible lentiviral plasmids.
