ABSTRACT
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ABSTRACT
Fusion of myoblasts into multinucleated myofibers is crucial for skeletal muscle development and regeneration. However, the mechanisms controlling this process remain to be determined. Here we identified the involvement of a new extracellular matrix protein in myoblast fusion. Collagen XXV is a transmembrane-type collagen highly transcribed during early myogenesis when primary myofibers form. Limb muscles of E12.5 and E14.5 Col25a1-/- embryos show a clear defect in the formation of multinucleated myofibers. In cell culture, the cleaved soluble extracellular domain of the collagen XXV is sufficient to promote the formation of highly multinucleated myofibers. Col25a1 is transiently expressed during myogenic differentiation and Col25a1 transcripts are down-regulated in multinucleated myofibers by a muscle-specific microRNA, miR-499. Altogether, these findings indicate that collagen XXV is required in vivo and in vitro for the fusion of myoblasts into myofibers and give further evidence that microRNAs participate to the regulation of this process.
Subject(s)
Cell Differentiation , Muscle Development , Non-Fibrillar Collagens/metabolism , 3' Untranslated Regions , Animals , Base Sequence , Cells, Cultured , Embryo, Mammalian/cytology , Embryo, Mammalian/metabolism , Humans , Mice , Mice, Knockout , MicroRNAs/chemistry , MicroRNAs/genetics , MicroRNAs/metabolism , Muscle, Skeletal/metabolism , Myoblasts/cytology , Myoblasts/metabolism , Non-Fibrillar Collagens/deficiency , Non-Fibrillar Collagens/genetics , Rats , Sequence AlignmentABSTRACT
Following injury, skeletal muscles can regenerate from muscle specific stem cells, called satellite cells. Quiescent in uninjured muscles, satellite cells become activated, proliferate and differentiate into myotubes. Muscle regeneration occurs following distinct main overlapping phases, including inflammation, regeneration and maturation of the regenerated myofibers. Each step of muscle regeneration is orchestrated through complex signaling networks and gene regulatory networks, leading to the expression of specific set of genes in each concerned cell type. Apart from the well-established transcriptional mechanisms involving the myogenic regulatory factors of the MyoD family, increasing data indicate that each step of muscle regeneration is controlled by a wide range of non-coding RNAs. In this review, we discuss the role of two classes of non-coding RNAs (microRNAs and long non-coding RNAs) in the inflammatory, regeneration and maturation steps of muscle regeneration.
ABSTRACT
Heat shock protein (HSP)72, the inducible form of HSP70, protects cells against a variety of injuries, but underlying mechanisms are poorly defined. To investigate the protective effects of HSP72, multiple clones expressing wild-type (WT) HSP72 and two mutants with defective nucleolar and nuclear localization (M45 and 985A, respectively) were made with the tet-off system in C2C12 cells. Four different parameters of cell function/injury were examined after simulated ischemia: protein synthesis, polysome formation, DNA synthesis, and lactate dehydrogenase (LDH release). Overexpression of WT HSP72 was also compared to nontransfected C2C12 cells. As expected, overexpression of HSP72 protected against simulated ischemia and reoxygenation for all parameters. In contrast, both M45 and 985A showed abnormal protein synthesis and polysome formation, both after simulated ischemia and under control conditions. Total RNA was slightly reduced in M45 and 985A at baseline, but 1 h after hypoxia, RNA levels were protected in all clones but significantly decreased in nontransfected C2C12 cells. Clones expressing 985A had nuclear retention of mRNA, suggesting that HSP72 is needed for nuclear export of RNA. All clones, both WT and mutant, had protection of DNA synthesis compared to C2C12 cells, but 985A had greater release of LDH after injury than any other group. These results support a multifactoral protective effect of HSP72, some aspects dependent on nuclear localization with stress and some not. The protection of protein synthesis and polysome formation, and abnormalities in these with the mutants, support a role for HSP72 in these processes both in the normal cell and in injury.
