Research Paper Volume 6, Issue 10 pp 820—834

Figure 1. SIRT-1 deactivates exercise-induced PARP-1 in skeletal muscle from young mice. Total RNA and cell lysates were isolated from the control or exercised gastrocnemius muscle. (A) Global cellular protein PARylation was determined in total cell lysates by immunoblots. PARP-1 (B) and SIRT-1 (C) mRNA (top) and protein (bottom) levels were determined in total muscle mRNA and cell lysate, respectively. GAPDH was used as a loading control. (D) PGC-1α acetylation levels were estimated by immunoblotting after IP. (E) mRNA expression of the indicated genes in the total RNA was examined by qPCR. (F) mtDNA was evaluated in total muscle genomic DNA by qPCR. (G) PARP-1 acetylation levels were estimated by immunoblotting after IP. (H) SIRT-1 and PARP-1 or (I) GCN5 and PARP-1 binding assays were estimated by immunoblotting after IP. (J) GCN5 protein levels were determined in total cell lysates by immunoblots. The blots are representative of three independent experiments. The data are presented as mean ± SEM (n = 3). White and black bars indicate non-exercised and exercised gastrocnemius muscle, respectively. COX17, cyclooxygenase 17; CytC, Cytochrome C; ERR-α, estrogen-related receptor α; mtTFA, mitochondrial transcription factor A; Ndufa2, NADH dehydrogenase [ubiquinone] iron-sulfur protein a 2; NRF1,nuclear respiratory factor 1; MCD, medium-chain acyl-CoA dehydrogenase; MCAD, medium-chain acyl-CoA dehydrogenase; SDH, succinate dehydrogenase; Tropn I, troponin I; UCP2, uncoupling protein 2; immunoprecipitation (IP).