Regionalisation of muscle nucleus heterogeneity in foetal skeletal muscle

Skeletal muscle is organized into functional domains with attachments at both extremities to tendons via the myotendinous junction (MTJ) and innervation in the middle via the neuromuscular junction (NMJ). In adult muscle, the molecular heterogeneity of muscle domains is described at the level of myonuclei with specialized MTJ and NMJ myonuclei at the tips and middle of muscle fibre. However, how molecular heterogeneity emerges during development remains poorly understood. Combining single-cell and single-nucleus RNA sequencing approaches with in situ hybridization experiments to limb muscles of chicken embryos, we uncover transcriptionally distinct muscle nuclei according to tip and central domains, both in myofibres and in mononucleated myogenic cells. At early foetal stage, all myonuclei display either a tip or a central domain identity. Immature mononucleated myogenic cells are enriched in the tip domain. Tip myonuclei of foetal muscles show a robust transcriptional signal transduction signature, which highlights molecular crosstalks between tip myonuclei and adjacent tendon fibroblasts but also with the adjacent immature mononucleated myogenic cells. We propose that the tip muscle domain is a signalling centre interacting with tendon fibroblasts to regulate muscle/tendon attachment and with adjacent mononucleated muscle cells to regulate myogenesis in limb foetal muscles.