Abstract
BackgroundMyogenin is a transcription factor that is expressed during terminal myoblast differentiation in embryonic development and adult muscle regeneration. Investigation of this cell state transition has been hampered by the lack of a sensitive reporter to dynamically track cells during differentiation.ResultsHere, we report a knock-in mouse line expressing the tdTOMATO fluorescent protein from the endogenous Myogenin locus. Expression of tdTOMATO in MyogntdTom mice recapitulated endogenous Myogenin expression during embryonic muscle formation and adult regeneration and enabled the isolation of the MYOGENIN+ cell population. We also show that tdTOMATO fluorescence allows tracking of differentiating myoblasts in vitro and by intravital imaging in vivo. Lastly, we monitored by live imaging the cell division dynamics of differentiating myoblasts in vitro and showed that a fraction of the MYOGENIN+ population can undergo one round of cell division, albeit at a much lower frequency than MYOGENIN− myoblasts.ConclusionsWe expect that this reporter mouse will be a valuable resource for researchers investigating skeletal muscle biology in developmental and adult contexts.
Highlights
Myogenin is a transcription factor that is expressed during terminal myoblast differentiation in embryonic development and adult muscle regeneration
Several reporter mouse lines have been generated to identify differentiating myoblasts based on the expression of Myosin light chain [25], Myog [26,27,28] and Muscle creatine kinase [29], they are based on lacZ or cat expression and only allow endpoint measurements on fixed samples
Generation and characterisation of a MyogntdTom mouse Using the Clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 system, a Single guide RNA (sgRNA) was designed to target the region of the STOP codon of the Myog gene for homologous recombination
Summary
Myogenin is a transcription factor that is expressed during terminal myoblast differentiation in embryonic development and adult muscle regeneration. Investigation of this cell state transition has been hampered by the lack of a sensitive reporter to dynamically track cells during differentiation. Embryonic and postnatal myogenesis and adult muscle regeneration are regulated by a family of basic helixloop-helix myogenic regulatory factors (MRFs) comprising Myf, Mrf, Myod and Myogenin (Myog). MuSCs activate the expression of Myod, proliferate to generate myoblasts that differentiate and fuse to form myofibres. Several reporter mouse lines have been generated to identify differentiating myoblasts based on the expression of Myosin light chain [25], Myog [26,27,28] and Muscle creatine kinase [29], they are based on lacZ (βgalactosidase activity, [30]) or cat (chloramphenicol acetyltransferase, [31]) expression and only allow endpoint measurements on fixed samples
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