Each skeletal muscle acquires its unique size before birth, when terminally differentiating myocytes fuse to form a defined number of multinucleated myofibres. Although mice in which the transcription factor Myogenin is mutated lack most myogenesis and die perinatally, a specific cell biological role for Myogenin has remained elusive. Here we report that loss of function of zebrafish myog prevents formation of almost all multinucleated muscle fibres. A second, Myogenin-independent, fusion pathway in the deep myotome requires Hedgehog signalling. Lack of Myogenin does not prevent terminal differentiation; the smaller myotome has a normal number of myocytes forming more mononuclear, thin, albeit functional, fast muscle fibres. Mechanistically, Myogenin binds to the myomaker promoter and is required for expression of myomaker and other genes essential for myocyte fusion. Adult myog mutants display reduced muscle mass, decreased fibre size and nucleation. Adult-derived myog mutant myocytes show persistent defective fusion ex vivo. Myogenin is therefore essential for muscle homeostasis, regulating myocyte fusion to determine both muscle fibre number and size.

Growth and maintenance of skeletal muscle fibres depend on coordinated activation and return to quiescence of resident muscle stem cells (MuSCs). The transcription factor Myogenin (Myog) regulates myocyte fusion during development, but its role in adult myogenesis remains unclear. In contrast to mice,myog^-/-zebrafish are viable, but have hypotrophic muscles. By isolating adult myofibres with associated MuSCs, we found thatmyog^-/-myofibres have severely reduced nuclear number, but increased myonuclear domain size. Expression of fusogenic genes is decreased, Pax7 upregulated, MuSCs are fivefold more numerous and mis-positioned throughout the length ofmyog^-/-myofibres instead of localising at myofibre ends as in wild-type. Loss of Myog dysregulates mTORC1 signalling, resulting in an ‘alerted’ state of MuSCs, which display precocious activation and faster cell cycle entry ex vivo, concomitant withmyodupregulation. Thus, beyond controlling myocyte fusion, Myog influences the MuSC:niche relationship, demonstrating a multi-level contribution to muscle homeostasis throughout life.

The Myocyte Enhancer Factor 2C (MEF2C) transcription factor plays a critical role in skeletal muscle differentiation, promoting muscle-specific gene transcription. Here we report that in proliferating cells MEF2C is degraded in mitosis by the Anaphase Promoting Complex/Cyclosome (APC/C) and that this downregulation is necessary for an efficient progression of the cell cycle. We show that this mechanism of degradation requires the presence on MEF2C of a D-box (R-X-X-L) and 2 phospho-motifs, pSer98 and pSer110. Both the D-box and pSer110 motifs are encoded by the ubiquitous alternate α1 exon. These two domains mediate the interaction between MEF2C and CDC20, a co-activator of APC/C. We further report that in myoblasts, MEF2C regulates the expression of G2/M checkpoint genes (14-3-3γ, Gadd45b and p21) and the sub-cellular localization of CYCLIN B1. The importance of controlling MEF2C levels during the cell cycle is reinforced by the observation that modulation of its expression affects the proliferation rate of colon cancer cells. Our findings show that beside the well-established role as pro-myogenic transcription factor, MEF2C can also function as a regulator of cell proliferation.

We describe molecular convergence between BMI1 and CHD7 in the initiation of medulloblastoma. Identified in a functional genomic screen in mouse models, a BMI1High;CHD7Low expression signature within medulloblastoma characterizes patients with poor overall survival. We show that BMI1-mediated repression of the ERK1/2 pathway leads to increased proliferation and tumor burden in primary human MB cells and in a xenograft model, respectively. We provide evidence that repression of the ERK inhibitor DUSP4 by BMI1 is dependent on a more accessible chromatin configuration in G4 MB cells with low CHD7 expression. These findings extend current knowledge of the role of BMI1 and CHD7 in medulloblastoma pathogenesis, and they raise the possibility that pharmacological targeting of BMI1 or ERK may be particularly indicated in a subgroup of MB with low expression levels of CHD7.

The transcription factor MEF2C (Myocyte Enhancer Factor 2C) plays an established role in the early steps of myogenic differentiation. However, the involvement of MEF2C in adult myogenesis and in muscle regeneration has not yet been systematically investigated. Alternative splicing of mammalian MEF2C transcripts gives rise to two mutually exclusive protein variants: MEF2Cα2 which exerts a positive control of myogenic differentiation, and MEF2Cα1, in which the α1 domain acts as trans-repressor of the MEF2C pro-differentiation activity itself. However, MEF2Cα1 variants are persistently expressed in differentiating cultured myocytes, suggesting a role in adult myogenesis. We found that overexpression of both MEF2Cα1/α2 proteins in a mouse model of muscle injury promotes muscle regeneration and hypertrophy, with each isoform promoting different stages of myogenesis. Besides the ability of MEF2Cα2 to increase differentiation, we found that overexpressed MEF2Cα1 enhances both proliferation and differentiation of primary myoblasts, and activates the AKT/mTOR/S6K anabolic signaling pathway in newly formed myofibers. The multiple activities of MEF2Cα1 are modulated by phosphorylation of Ser98 and Ser110, two amino acid residues located in the α1 domain of MEF2Cα1. These specific phosphorylations allow the interaction of MEF2Cα1 with the peptidyl-prolyl isomerase PIN1, a regulator of MEF2C functions. Overall, in this study we established a novel regulatory mechanism in which the expression and the phosphorylation of MEF2Cα1 are critically required to sustain the adult myogenesis. The described molecular mechanism will represent a new potential target for the development of therapeutical strategies to treat muscle-wasting diseases.

Deregulation of chromatin modifiers plays an essential role in the pathogenesis of medulloblastoma, the most common paediatric malignant brain tumour. Here, we identify a BMI1-dependent sensitivity to deregulation of inositol metabolism in a proportion of medulloblastoma. We demonstrate mTOR pathway activation and metabolic adaptation specifically in medulloblastoma of the molecular subgroup G4 characterised by a BMI1^High;CHD7^Low signature and show this can be counteracted by IP6 treatment. Finally, we demonstrate that IP6 synergises with cisplatin to enhance its cytotoxicity in vitro and extends survival in a pre-clinical BMI1^High;CHD7^Low xenograft model.