Title

 

Authors

C. De Palma¹; S. Pisoni¹; S. Pambianco¹; S. Falcone⁴; L. Scorrano²; M. Sandri³ E. Clementi<sup>1

1</sup>Unit of Clinical Pharmacology,  Dep. of Clinical Sciences, "Luigi Sacco" University Hospital of Milano, Italy;²Department of Cell Physiology and Metabolism, University of Geneva, 1211 Geneva, Switzerland;³ VIMM, Venetian Institute of Molecular Medicine, Padua, Italy; ⁴ UMR S 787 Inserm Institute de Myologie, Paris

 

Abstract

Mitochondrial morphology is regulated by the balanced processes of fission and fusion; whether these events play a role also in shaping cell functions is still not well understood. Here we provide evidence that during myogenic differentiation the short mitochondria of myoblasts change into the extensively elongated network observed in myotubes. We show that mitochondrial elongation is required for myogenesis to occur and that this event depends on the cellular generation of nitric oxide (NO). Inhibition of NO synthesis in myogenic precursor cells leads to inhibition of mitochondrial elongation and of myogenic differentiation. The specific target of NO action is Drp1. NO controls Drp1 translocation to mitochondria and its docking to the specific interactor Fis1; further, it controls the GTPase activity of the protein. In the presence of NO Drp1 functions are maintained at basal levels to allow mitochondria to create a fused network that enhances myogenic differentiation. The action of NO on Drp1 is mediated through a direct phosphorylation of the protein, induced by activation of PKG. Inhibition of the NO pathway leads to mitochondrial fission  accounting for a latent mitochondrial dysfunction, reducing respiratory activity and OXPHOS ATP production. Despite the apoptotic features of these mitochondria, myoblasts do not undergo spontaneous apoptosis. They acquire sensitivity to apoptosis in the presence of 3-methyl-adenine, a general inhibitor of lysosomes degradation. In addition we found a decrease of mitochondrial number and DNA when NO generation was inhibited, accompanied by increases in LC3 positive autophagosomes and cleveage of LC3. These findings suggest that mitochondrial latent dysfunction is related to induction of a mitophagic prosurvival pathway.
In this study we demonstrate a new regulation of Drp1 function mediated by NO and the fundamental role played by this regulation during myogenic differentiation. In addition we show how mitophagy can act as a protective factor in the absence of NO. These results unravel a fine  control check of the myogenic differentiation programme dependent on metabolic signals.