(A-C) HEK293T cells were transfected with either scramble (control) or PARP-1 shRNA and HA-PGC-1α expression vector for 48h. Then, (A) PARP-1 protein levels and autoPARylation (arrowhead) were analyzed in total protein lysates. (B) Intracellular NAD+ was measured on total acid extracts. (C) PGC-1α acetylation was analyzed in HA immunoprecipitates. (D-F) HEK293T cells were transfected with either a pool of PARP-1 siRNAs, a pool of SIRT1 siRNAs, or different combinations of both using the corresponding scramble siRNAs as control (−). The cells were simultaneously transfected with HA-PGC-1α for 48h. Then, (D) relative mitochondrial DNA content, (E) mRNA levels of the genes indicated and (F) total O2 consumption was analyzed. * indicates statistical difference vs. respective control sh/siRNA-transfected cells at p<0.05.