NuMA1 controls myonuclear motility in striated skeletal muscle through AMPK activity and is impaired in Duchenne Muscular Dystrophy

Skeletal muscle consists in a bundle of thousands post-mitotic multinucleated cells, called myofibers, in which myonuclei are evenly spaced and positioned at the periphery. This myonuclear positioning i) shapes myonuclear domain (MND) in myofibers, essential for the transcriptional integrity of myofibers, ii) is driven by cytoskeleton and associated proteins and iii) is required for proper myofiber functions. In numerous of muscle diseases (i.e. myopathies), alteration of myonuclei localization (internalized and/or mispositioned) contributes to myofiber misfunctioning, supporting the need to better understand the fundamental mechanisms that regulates myonuclei dynamics and to restore the establishment of myonuclear domains in these diseases. In this study, we show that in Duchenne muscular dystrophy (DMD) myofibers, myonuclei are more dynamic and contribute to the failure in MNDs settings, suggesting that in pathological conditions enhanced myonuclear motility impact myonuclear distribution. To identify new actors regulating this MND settings, we performed a mass spectrometry (MS)-based proteomic analysis to identify microtubule associated proteins (MAPs) in myotubes/myofibers and performed a siRNA screening on selected candidates. This approach allowed to highlight NuMA1 as a new factor controlling myoblast fusion and myonuclear positioning through the control of nuclear-microtubule-organizing-center (n-MTOC) integrity and microtubule network orientation. Strikingly, while NuMA1 is restrained to myonuclei in mononucleated myoblasts, it progressively accumulates in the cytoplasm during muscle cell differentiation, preferentially with microtubule (MT) nucleation spots at the vicinity of the nuclear membrane. We identified that AMP Kinase activity has an essential role in NuMA1 nuclear accumulation through the specific phosphorylation on serine-1853 and the ability of myonuclei to accumulate NuMA1 is correlated to their motility in myofibers. Finally, we show that nuclear NuMA1 content is increased in DMD patients and mdx mouse model, contributing to more dynamic myonuclei that can manipulated pharmacologically through the control of AMPK activity. Altogether, our data identify a novel mechanism by which nuclear sequestration of a MAP allows to couple nuclear positioning and motion to MT organization.