Incorporation of a skeletal muscle-specific enhancer in the regulatory region of Igf1 upregulates IGF1 expression and induces skeletal muscle hypertrophy

Abstract

Abstract In this study, we upregulated insulin-like growth factor-1 (IGF1) expression specifically in skeletal muscle by engineering an enhancer into its non-coding regions and verified the expected phenotype in a mouse model. To select an appropriate site for introducing a skeletal muscle-specific myosin light chain (MLC) enhancer, three candidate sites that exhibited the least evolutionary conservation were chosen and validated in C2C12 single-cell colonies harbouring the MLC enhancer at each site. IGF1 was dramatically upregulated in only the site 2 single-cell colony series, and it exhibited functional activity leading to the formation of extra myotubes. Therefore, we chose site 2 to generate a genetically modified (GM) mouse model with the MLC enhancer incorporated by CRISPR/Cas9 technology. The GM mice exhibited dramatically elevated IGF1 levels, which stimulated downstream pathways in skeletal muscle. Female GM mice exhibited more conspicuous muscle hypertrophy than male GM mice. The GM mice possessed similar circulating IGF1 levels and tibia length as their WT littermates; they also did not exhibit heart abnormalities. Our findings demonstrate that genetically modifying a non-coding region is a feasible method to upregulate gene expression and obtain animals with desirable traits.