MicroRNAs (miRNAs) are a class of small non-coding RNAs that have recently emerged as important regulators of gene expression. They negatively regulate gene expression post-transcriptionally by translational repression and target mRNA degradation. miRNAs have been shown to play crucial roles in muscle development and in regulation of muscle cell proliferation and differentiation. By comparing miRNA expression profiling of proliferating myoblasts versus differentiated myotubes, a number of modulated miRNAs, not previously implicated in regulation of myogenic differentiation, were identified. Among these, miR-221 and miR-222 were strongly down-regulated upon differentiation of both primary and established myogenic cells. Conversely, miR-221 and miR-222 expression was restored in post-mitotic, terminally differentiated myotubes subjected to Src tyrosine kinase activation. By the use of specific inhibitors we provide evidence that expression of miR-221 and miR-222 is under the control of the Ras-MAPK pathway. Both in myoblasts and in myotubes, levels of the cell cycle inhibitor p27 inversely correlated with miR-221 and miR-222 expression, and indeed we show that p27 mRNA is a direct target of these miRNAs in myogenic cells. Ectopic expression of miR-221 and miR-222 in myoblasts undergoing differentiation induced a delay in withdrawal from the cell cycle and in myogenin expression, followed by inhibition of sarcomeric protein accumulation. When miR-221 and miR-222 were expressed in myotubes undergoing maturation, a profound alteration of myofibrillar organization was observed. miR-221 and miR-222 have been found to be modulated during myogenesis and to play a role both in the progression from myoblasts to myocytes and in the achievement of the fully differentiated phenotype. Identification of miRNAs modulating muscle gene expression is crucial for the understanding of the circuits controlling skeletal muscle differentiation and maintenance.

A class of small non-coding RNAs, the microRNAs (miRNAs), have recently attracted great attention in cancer research since they play a central role in regulation of gene-expression and miRNA aberrant expression is found in almost all types of human cancer. The discovery of circulating miRNAs in body fluids and the finding that they are often tumor specific and can be detected early in tumorigenesis has soon led to the evaluation of their possible use as cancer biomarkers and treatment-response predictors. The evidence that tumor cells communicate via the secretion and delivery of miRNAs packed into tumor-released microvesicles has prompted to investigate miRNA contribution as signaling molecules to the establishment and maintenance of the tumor microenvironment and the metastatic niche in cancer. In this review we highlight the recent advances on the role of exosomal miRNAs as mediators of cancer cell-to-cell communication.

The relationship between susceptibility to transformation in vitro by different oncogenes and terminal differentiation was analyzed in embryonic quail myogenic cells. Infection with Rous sarcoma virus (RSV), Fujinami sarcoma virus (FSV), avian erythroblastosis virus (AEV), and the avian myelocytomatosis virus MC29 led to rapid and massive transformation. Transformed cells had distinctive morphological alterations, increased proliferation rates, and the ability to grow in agar suspension. Furthermore, homogeneously transformed cultures failed to fuse into multinucleated myotubes and to express muscle-specific genes. However, cloned populations of RSV-, FSV-, and AEV-transformed myogenic cells could, under appropriate culture conditions, partially differentiate into atypical "revertant" myotubes. In contrast, competence for terminal differentiation was completely and irreversibly suppressed on transformation by MC29. The specificity of action of a given oncogenic sequence on the inhibition of differentiation was further studied by using conditional and nonconditional transformation mutants. Myogenic cells infected with temperature-sensitive (ts) mutants of RSV and FSV exhibited a fully reversible block of differentiation after shift to restrictive temperature, while cells infected with ts34 AEV were not temperature sensitive for differentiation. Cultures infected with the partially transformation-defective mutant of MC29 td10H were morphologically transformed and acquired anchorage independence for proliferation but maintained a residual competence for terminal differentiation.

Circular RNAs (circRNAs) are a class of RNA produced during pre-mRNA splicing that are emerging as new members of the gene regulatory network. In addition to being spliced in a linear fashion, exons of pre-mRNAs can be circularized by use of the 3′ acceptor splice site of upstream exons, leading to the formation of circular RNA species. In this way, genetic information can be re-organized, increasing gene expression potential. Expression of circRNAs is developmentally regulated, tissue and cell-type specific, and shared across eukaryotes. The importance of circRNAs in gene regulation is now beginning to be recognized and some putative functions have been assigned to them, such as the sequestration of microRNAs or proteins, the modulation of transcription, the interference with splicing, and translation of small proteins. In accordance with an important role in normal cell biology, circRNA deregulation has been reported to be associated with diseases. Recent evidence demonstrated that circRNAs are highly expressed in striated muscle tissue, both skeletal and cardiac, that is also one of the body tissue showing the highest levels of alternative splicing. Moreover, initial studies revealed altered circRNA expression in diseases involving striated muscle, suggesting important functions of these molecules in the pathogenetic mechanisms of both heart and skeletal muscle diseases. The recent findings in this field will be described and discussed.

Myotonic dystrophy type 1 (DM1) is the most common adult-onset muscular dystrophy, characterized by progressive myopathy, myotonia, and multi-organ involvement. This dystrophy is an inherited autosomal dominant disease caused by a (CTG)n expansion within the 3' untranslated region of the DMPK gene. Expression of the mutated gene results in production of toxic transcripts that aggregate as nuclear foci and sequester RNA-binding proteins, resulting in mis-splicing of several transcripts, defective translation, and microRNA dysregulation. No effective therapy is yet available for treatment of the disease. In this study, myogenic cell models were generated from myotonic dystrophy patient-derived fibroblasts. These cells exhibit typical disease-associated ribonuclear aggregates, containing CUG repeats and muscleblind-like 1 protein, and alternative splicing alterations. We exploited these cell models to develop new gene therapy strategies aimed at eliminating the toxic mutant repeats. Using the CRISPR/Cas9 gene-editing system, the repeat expansions were removed, therefore preventing nuclear foci formation and splicing alterations. Compared with the previously reported strategies of inhibition/degradation of CUG expanded transcripts by various techniques, the advantage of this approach is that affected cells can be permanently reverted to a normal phenotype.

The RhoA GTPase controls a variety of cell functions such as cell motility, cell growth, and gene expression. Previous studies suggested that RhoA mediates signaling inputs that promote skeletal myogenic differentiation. We show here that levels and activity of RhoA protein are down-regulated in both primary avian myoblasts and mouse satellite cells undergoing differentiation, suggesting that a fine regulation of this GTPase is required. In addition, ectopic expression of activated RhoA in primary quail myocytes, but not in mouse myocytes, inhibits accumulation of muscle-specific proteins and cell fusion. By disrupting RhoA signaling with specific inhibitors, we have shown that this GTPase, although required for cell identity in proliferating myoblasts, is not essential for commitment to terminal differentiation and muscle gene expression. Ectopic expression of an activated form of its downstream effector, Rock, impairs differentiation of both avian and mouse myoblasts. Conversely, Rock inhibition with specific inhibitors and small interfering RNA-mediated gene silencing leads to accelerated progression in the lineage and enhanced cell fusion, underscoring a negative regulatory function of Rock in myogenesis. Finally, we have reported that Rock acts independently from RhoA in preventing myoblast exit from the cell cycle and commitment to differentiation and may receive signaling inputs from Raf-1 kinase.