SH3KBP1 scaffolds endoplasmic reticulum and controls skeletal myofibers architecture and integrity

Abstract

The building block of skeletal muscle is the multinucleated muscle fiber, formed by the fusion of hundreds of mononucleated precursor cells, myoblasts. In the normal course of muscle fiber development or regeneration, myonuclei are actively positioned throughout muscular development and adopt special localization in mature fibers: regular spacing along muscle fibers periphery, raising the notion of MyoNuclear Domains (MNDs). There is now growing support for a direct connection between myonuclear positioning and normal function of muscles, but how myonuclei affects muscle function remains poorly characterized. To identify new factors regulating forces applied on myonuclei in muscles fibers, we performed a siRNA screen and identified SH3KBP1 as a new factor controlling myonuclear positioning in early phases of myofibers formation. Depletion of SH3KBP1 induces a reset of MNDs establishment in mature fibers reflected by a dramatic reduction in pairwise distance between myonuclei. We show that SH3KBP1 scaffolds Endoplasmic Reticulum (ER) in myotubes that in turn controls myonuclei velocity and localization and thus myonuclear domains settings. Additionally, we show that in later phases of muscle maturation, SH3KBP1 contributes to the formation and maintenance of Sarcoplasmic Reticulum (SR) and Transverse-tubules (T-tubules). We also demonstrate that in muscle fibers, GTPase dynamin-2 (DNM2) binds to SH3 domains of SH3KBP1. Interestingly, we observed that Sh3kbp1 mRNA is up regulated in a mouse model harboring the most frequent mutation for Autosomal Dominant CentroNuclear Myopathy (AD-CNM): Dnm2+/R465W. SH3KBP1 thus appears as a compensation mechanism in this CNM model since its depletion contributes to an increase of CNM-like phenotypes (reduction of muscle fibers Cross-section Areas (CSA) and increase in slow fibers content). Altogether our results identify SH3KBP1 as a new regulator of myonuclear domains establishment in the early phase of muscle fibers formation through ER scaffolding and later in myofibers integrity through T-tubules scaffolding/maintenance. SummaryMyonuclei are actively positioned throughout muscular development. Guiraud, Christin, Couturier et al show that SH3KBP1 scaffolds the ER through Calnexin interaction and controls myonuclei motion during early steps of muscle fibers formation. Besides SH3KBP1 participates in cell fusion and T-tubules formation/maintenance in mature skeletal muscle fibers and contributes to slow-down CNM-like phenotypes.