Microtubule-dependent transport and organization of sarcomeric myosin during skeletal muscle differentiation

Abstract

It has been proposed that microtubules (MTs) participate in skeletal muscle cell differentiation. However, it is still unclear how this happens. To examine whether MTs could participate directly in the organization of thick and thin filaments into sarcomeres, we observed the concomitant reorganization and dynamics of MTs with the behavior of sarcomeric actin and myosin by time-lapse confocal microscopy. Using green fluorescent protein (GFP)-EB1 protein to label MT plus ends, we determined that MTs become organized into antiparallel arrays along fusing myotubes. Their dynamics and orientation was found to be different across the thickness of the myotubes. We observed fast movements of Dsred-myosin along GFP-MTs. Comparison of GFP-EB1 and Dsred-myosin dynamics revealed that myosin moved toward MT plus ends. Immuno-electron microscopy experiments confirmed that myosin was actually associated with MTs in myotubes. Finally, we confirmed that MTs were required for the stabilization of myosin-containing elements prior to incorporation into mature sarcomeres. Collectively, our results strongly suggest that MTs become organized into a scaffold that provides directional cues for the positioning and organization of myosin filaments during sarcomere formation.

Solute carrier family 3 member 2 (Slc3a2) controls yolk syncytial layer (YSL) formation by regulating microtubule networks in the zebrafish embryo

“…5). Realignment of microtubules into parallel arrays along the axis of the fusing cells has been shown to be crucial for syncytial myotube formation (36)(37)(38). These data support the view that microtubules control the spatiotemporal aspects of plasma membrane fusion in YSL progenitors.…”

Myoseverin disrupts sarcomeric organization in myocytes: An effect independent of microtubule assembly inhibition

“…Our results contradict the previous suggestion that active trafficking of myosin along MTs is essential for sarcomere assembly in skeletal myocytes [Pizon et al, 2005]. The possibility remains that MT functions differ between skeletal and cardiac myocytes as there are subtle differences in sarcomere assembly between muscle types [Lloyd et al, 2004].…”

“…The possibility remains that MT functions differ between skeletal and cardiac myocytes as there are subtle differences in sarcomere assembly between muscle types [Lloyd et al, 2004]. Furthermore, Pizon and colleagues make the distinction that nocodazole induced MT disassembly only affected new sarcomere assembly as nascent myofibrils but not mature myofibrils were lost [Pizon et al, 2005]. In our cardiac myocyte model, no mature myofibrils were observed in non-hypertrophic control cells as titin staining (Fig.…”

“…More recently, it has also been proposed that sarcomeric myosin is transported along MTs and directed to the site of assembly during myofibrillogenesis in skeletal muscle [Pizon et al, 2005]. In contrast, a recent study has indicated that MTs could not serve as scaffolds for sarcomere formation [Siebrands et al, 2004].…”

Generation of noncentrosomal microtubule arrays

“…Notably, many MTs in the array become stabilized as the cells differentiate (Gundersen et al, 1989). The linear, noncentrosomal array of MTs in myotubes has been implicated in myofibrillogenesis as well as in establishing the elongated shape of the cell (Antin et al, 1981;Hill et al, 1986;Pizon et al, 2005;Tassin et al, 1985). Experiments using inhibitors of MTs have also implicated MTs in the mobility of acetylcholine receptors in primary cultures of chick embryonic muscle cells, suggesting a function for these arrays in the development of neuromuscular junctions (Connolly, 1984;Connolly and Oldfin, 1985).…”