Myonuclear domain and microtubule proteome during skeletal muscle maturation

In the normal course of muscle fiber development, myonuclei actively position and adapt a precise localization in mature fibers, shaping MyoNuclear Domains (MNDs). Myonuclei positioning in fibers appears to be essential for muscle function as defects in MNDs settings are always associated with dysfunction (i.e., centronuclear myopathy, sarcopenia). Previous studies have shown that myonuclei positioning in fibers is reversible, suggesting that in pathologies presenting MNDs impairment, myonuclei could be re-addressed to the "correct" position in fibers and this could benefit to muscle function. Cytoskeleton networks, and particularly microtubules, have been implicated in early nuclei localization in myotubes. As the microtubule network is completely redesigned during muscle maturation, we hypothesized that "microtubules associated proteomes" would change between immature and mature fibers and contribute to a microtubule-dependent process resulting in MNDs setting and maintenance in mature fibers. We performed an in vitro biochemical approach to isolate microtubules partners in immature (myotubes) and mature myofibers. Using mass-spectrometry identification, we selected 244 candidates, differentially associated/expressed with microtubules during myofiber maturation and potentially controlling MNDs settings. We are currently conducting a siRNA screen approach on these candidates to decipher their respective implication in early and late phases of MNDs establishment, using an unbiased assay developed by our team allowing statistical analysis of MNDs regarding myonuclei content. This approach will lead to the identification of new pathways related to nuclear positioning and MNDs setting in normal condition and in myopathies associated to MNDs impairment such as CNMs.