Abstract
Palladin is a microfilament-associated phosphoprotein whose function in skeletal muscle has rarely been studied. Therefore, we investigate whether myogenesis is influenced by the depletion of palladin expression known to interfere with the actin cytoskeleton dynamic required for skeletal muscle differentiation. The inhibition of palladin in C2C12 myoblasts leads to precocious myogenic differentiation with a concomitant reduction in cell apoptosis. This premature myogenesis is caused, in part, by an accelerated induction of p21, myogenin, and myosin heavy chain, suggesting that palladin acts as a negative regulator in early differentiation phases. Paradoxically, palladin-knockdown myoblasts are unable to differentiate terminally, despite their ability to perform some initial steps of differentiation. Cells with attenuated palladin expression form thinner myotubes with fewer myonuclei compared to those of the control. It is noteworthy that a negative regulator of myogenesis, myostatin, is activated in palladin-deficient myotubes, suggesting the palladin-mediated impairment of late-stage myogenesis. Additionally, overexpression of 140-kDa palladin inhibits myoblast differentiation while 200-kDa and 90-kDa palladin-overexpressed cells display an enhanced differentiation rate. Together, our data suggest that palladin might have both positive and negative roles in maintaining the proper skeletal myogenic differentiation in vitro.
Highlights
Skeletal muscle differentiation is a highly ordered multiphase process from myoblast proliferation, to fusion, to myotube differentiation [1,2,3]
Results short hairpin RNA (shRNA)-mediated knockdown of palladin accelerates the onset of C2C12 myoblast differentiation
In order to provide a dynamic analysis of myoblast differentiation into multinucleated myotubes, the myogenic index was assessed by counting MyHCpositive cells (Fig 1E, arrows)
Summary
Skeletal muscle differentiation is a highly ordered multiphase process from myoblast proliferation, to fusion, to myotube differentiation [1,2,3]. Dual Functions of Palladin in Skeletal Muscle Differentiation facilitates cell fusion and commits myoblasts to withdraw from the cell cycle [7]. Myoblasts still appear mononucleated but irreversibly withdraw from the cell cycle. In this phase, a portion of undifferentiated or partly differentiated cells undergoes apoptosis [10]. In late myogenic differentiation events, myotubes undergo further maturation to generate functional muscle cells, as evidenced by increases in size and changes in the expression of contractile proteins [7, 11, 12]. The sub-cellular coordination of the cytoskeleton and its regulatory, scaffolding, and cytoskeletal cross-linking proteins are responsible for reorganizations and maintaining the normal actin cytoskeleton during myogenesis [14,15,16]
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