Abstract
Skeletal muscles are essential for movement, supporting a wide range of locomotor behaviors. Muscle tissue is composed of multiple cell types including “fast” and “slow” myofibers, whose contractile properties are largely influenced by selective expression of myosin heavy chain (MyHC) isoforms. While ‘super-enhancers’ regulating MyHC gene clusters have been identified, the cis-regulatory elements (CREs) controlling non-MyHC genes important to myofiber physiology remain less defined. Here, we profile the regulatory landscape of two pairs of mouse hind limb muscles differing in MyHC expression at a late embryonic (E18.5) and adult time point to identify candidate CREs that may regulate genes important to myofiber type. Gene expression and chromatin accessibility analyses revealed that epigenetic differences at E18.5 largely reflect limb patterning, whereas adult differences reflect myofiber differentiation. We identified thousands of differentially accessible regions that may regulate genes important for muscle development, muscle biology, and myofiber identity. Among these, twelve conserved, muscle-specific CREs associated with myofiber type were tested for regulatory activity. Nine enhanced and three reduced gene activity in vitro, although their phenotypic effects remain unknown. By profiling multiple muscles across two time points, our study extends current understanding of conserved, muscle-specific CREs that regulate gene expression during myogenesis.
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Data availability
All ATAC-seq and RNA-seq datasets generated as part of this study have been deposited to the NCBI GEO repository under accession number GSE292230 and GSE292232. All data included in this study are available upon reasonable request by contact with the corresponding author.
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Acknowledgements
This study was supported by the University of Oregon, National Science Foundation (SRQ BCS-1945809 and ASO DGE-1745303) and the Leakey Foundation. Thanks to Emmanuelle Boucicaut, Joseph Braud, Jack Chambers, Emma Freedman and Elijah Reed for their help in the cell culture lab and to Natalie Dunn, Carrie McCurdy, Kristin Kohler, Daniel Richard, Avika Gomez-Sharma, Allissa Van Steenis and Mariel Young for their expertise and technical support. This work benefited from access to the University of Oregon high performance computing cluster, Talapas. Some figures were created using BioRender.
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SRQ, TDC, and KNS conceptualized the study; KNS and TDC supervised the project; ASO and SRQ performed the experiments; SRQ wrote the manuscript, analyzed the data, and prepared the figures; MCO and DMC substantially revised the manuscript for intellectual content, and all authors edited the manuscript. All authors approved the final version of the manuscript.
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Queeno, S.R., Okamoto, A.S., Callahan, D.M. et al. Profiling the epigenomic landscape of late embryonic and adult mouse hind limb muscles. Sci Rep (2026). https://doi.org/10.1038/s41598-025-32705-4
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DOI: https://doi.org/10.1038/s41598-025-32705-4
