Tension-driven multi-scale self-organisation in human iPSC-derived muscle fibers

Abstract

Human muscle is a hierarchically organised tissue with its contractile cells called myofibers packed into large myofiber bundles. Each myofiber contains periodic myofibrils built by hundreds of contractile sarcomeres that generate large mechanical forces. To better understand the mechanisms that coordinate human muscle morphogenesis from tissue to molecular scales, we adopted a simple in vitro system using induced pluripotent stem cell-derived human myogenic precursors. When grown on an unrestricted two-dimensional substrate, developing myofibers spontaneously align and self-organise into higher-order myofiber bundles, which grow and consolidate to stable sizes. Following a transcriptional boost of sarcomeric components, myofibrils assemble into chains of periodic sarcomeres that emerge across the entire myofiber. More efficient myofiber bundling accelerates the speed of sarcomerogenesis suggesting that tension generated by bundling promotes sarcomerogenesis. We tested this hypothesis by directly probing tension and found that tension build-up precedes sarcomere assembly and increases within each assembling myofibril. Furthermore, we found that myofiber ends stably attach to other myofibers using integrin-based attachments and thus myofiber bundling coincides with stable myofiber bundle attachment in vitro. A failure in stable myofiber attachment results in a collapse of the myofibrils. Overall, our results strongly suggest that mechanical tension across sarcomeric components as well as between differentiating myofibers is key to coordinate the multi-scale self-organisation of muscle morphogenesis.

Stiffness anisotropy coordinates supracellular contractility driving long-range myotube-ECM alignment

“…Following mLCN film preparation, we seeded C2C12 myoblasts at a relatively low density (5000 cells/cm 2 ) on sterilized mLCNs as well as gelatin-coated glass coverslips for an additional isotropic, stiff control used in myoblast culture ( 24 ). Consistent with previous reports, we observed that C2C12s adhered to neat mLCNs, presumably due to nonspecific protein adsorption from the growth medium ( 13 , 16 , 25 ).…”

Cellular Dynamics of Skeletal Muscle Regeneration

“…Instead, they arise after fusion and are strongly correlated with myofibril assembly, suggesting that positioning of myonuclei is regulated by myofibrillogenesis. Previous studies have proposed a link between myofibril formation and myonuclei location 67,79,80 , but in vitro studies have found that forming myofibrils drive nuclei to the myofiber periphery 67 . However, in myofibers regenerated in vivo after a catastrophic injury, the initial myonuclei remain in centralized chains; we hypothesize that the rapidity of myocyte-myocyte fusion, nearly synchronous formation of myofibrils, and the enclosing BM traps and positions myonuclei into centralized chains.…”

Styxl2 regulates de novo sarcomere assembly by binding to non-muscle myosin IIs and promoting their degradation

“…Each of these models is supported by some experimental observations but may be incompatible with others ( Chopra et al, 2018 ; Fenix et al, 2018 ; Rui et al, 2010 ). In addition, mechanical tension generated by attachment of myofibers to tendon, myofiber bundling, and formation of mini-sarcomeres is also found to promote sarcomere assembly in both fly and mammalian muscle cells ( Chopra et al, 2018 ; Loison et al, 2018 ; Luis and Schnorrer, 2021 ; Mao et al, 2022 ; Weitkunat et al, 2014 ).…”