br Experimental section br Competing interests br Author s

Experimental section
Competing interests
Author's contributions
Acknowledgement This research is partially supported by a seed fund for the Center for Drug Discovery and Translational Research (LS) and a research grant by Fujifilm (VPS and LS). The Sponsors played no roles in study design, the collection, analysis and interpretation of data, the writing of the report, and the decision to submit the article for publication. We thank Charles Sheehan of the NMR Core of Harvard Medical School for his critical advice on 13C-HMBC/13C-HSQC experiments and structural determinations of compound 2c, and BPS Bioscience for technical support in the ADP Glo enzymatic assay.
Introduction Skeletal muscle displays a high ability to regenerate due to the presence of satellite Concanamycin A (SCs) (Blau et al., 2015). Myofiber formation from quiescent SCs follows a highly coordinated multi-step process orchestrated by a number of muscle regulatory factors (MRFs) (Tapscott, 2005). Upon injury, quiescent SCs, characterized by high Pax7 expression, become proliferating myoblasts with induced expression of the basic helix-loop-helix myogenic regulatory factors Myf5 and MyoD (Sartorelli and Caretti, 2005). Myogenin, the other member of the basic helix-loop-helix myogenic regulatory factors, is highly expressed in the early phase of differentiation (Sartorelli and Caretti, 2005). The myoblasts then fuse into fully differentiated myofibers, characterized by myosin expression (MyHC), among other markers (Wang et al., 2014). MRF expression is controlled at the transcriptional level (Brack and Rando, 2012), however, signaling pathways and chromatin remodeling further regulate their expression (Segalés et al., 2016). In this context, the role of histone acetylases (HATs) and histone deacetylases (HDACs) has been reported to modulate the expression of MRFs, thus influencing muscle differentiation (Polesskaya et al., 2001, Sincennes et al., 2016). Importantly, p300 acetyl transferase activity has been shown to be critical for the regulation of Myod expression and skeletal muscle development (Roth et al., 2003, Hamed et al., 2013). Also, differentiation signals induce a change in the NAD+/NADH ratio leading to Sirt2 inhibition, which in turn increases histone acetylation (Sartorelli and Caretti, 2005). We recently reported that IGF-1-induced phosphorylation and activation of ATP citrate lyase (ACL) through the PI3K/AKT pathway regulate mitochondrial function, as well as glucose and lipid metabolism in skeletal muscle (Das et al., 2015). ACL catalyzes the conversion of citrate into oxaloacetate and acetyl-CoA. Acetyl-CoA is further utilized by histone acetylases to acetylate histones, and regulation of histone acetylation by ACL has been reported in proliferating cancer cells (Covarrubias et al., 2016, Lee et al., 2014, Wellen et al., 2009, Zhao et al., 2016). Intrigued by the prominent role ACL has in regulating histone acetylation, we investigated whether ACL regulates SC proliferation and differentiation and, finally, muscle regeneration.
Results
Discussion We recently demonstrated that ACL regulates mitochondrial function by controlling levels of the critical mitochondrial lipid cardiolipin. Activation of the IGF-1/Akt/ACL/cardiolipin pathway is sufficient to increase mitochondrial supercomplex formation, resulting in increases in ATP (Das et al., 2015). In addition to this vital function, ACL catalyzes a critical step in the formation of acetyl-CoA, which is further utilized by histone acetylases to acetylate histones. What wasn’t clear was whether acetyl-CoA was limiting or whether driving the formation of more acetyl-CoA was sufficient to perturb patterns of histone acetylation, regulating transcription in skeletal muscle, as described in cancer cells (Covarrubias et al., 2016, Lee et al., 2014, Wellen et al., 2009, Zhao et al., 2016). Given the prominent role of histone acetylation in regulating the expression of MRFs (Polesskaya et al., 2001, Sincennes et al., 2016), we investigated whether ACL regulates skeletal muscle differentiation and regeneration.