Deep Sequencing the microRNA profile in rhabdomyosarcoma reveals down-regulation of miR-378 family members
Rhabdomyosarcoma (RMS) is a highly malignant tumour accounting for nearly half of soft tissue sarcomas in children. MicroRNAs (miRNAs) represent a class of short, non-coding, regulatory RNAs which play a critical role in different cellular processes.
Altered miRNA levels have been reported in human cancers, including RMS.
Methods: Using deep sequencing technology, a total of 685 miRNAs were investigated in a group of alveolar RMSs (ARMSs), embryonal RMSs (ERMSs) as well as in normal skeletal muscle (NSM). Q-PCR, MTT, cytofluorimetry, migration assay, western blot and immunofluorescence experiments were carried out to determine the role of miR-378a-3p in cancer cell growth, apoptosis, migration and differentiation.
Bioinformatics pipelines were used for miRNA target prediction and clustering analysis.
Results: Ninety-seven miRNAs were significantly deregulated in ARMS and ERMS when compared to NSM. MiR-378 family members were dramatically decreased in RMS tumour tissue and cell lines.
Interestingly, members of the miR-378 family presented as a possible target the insulin-like growth factor receptor 1 (IGF1R), a key signalling molecule in RMS. MiR-378a-3p over-expression in an RMS-derived cell line suppressed IGF1R expression and affected phosphorylated-Akt protein levels.
Ectopic expression of miR-378a-3p caused significant changes in apoptosis, cell migration, cytoskeleton organization as well as a modulation of the muscular markers MyoD1, MyoR, desmin and MyHC. In addition, DNA demethylation by 5-aza-2[prime]-deoxycytidine (5-aza-dC) was able to up-regulate miR-378a-3p levels with a concomitant induction of apoptosis, decrease in cell viability and cell cycle arrest in G2-phase.
Cells treated with 5-aza-dC clearly changed their morphology and expressed moderate levels of MyHC.
Conclusions: MiR-378a-3p may function as a tumour suppressor in RMS and the restoration of its expression would be of therapeutic benefit in RMS. Furthermore, the role of epigenetic modifications in RMS deserves further investigations.
Author: Francesca MegiorniSamantha CialfiHeather P McDowellArmando FelsaniSimona CameroAlessandro GuffantiBarry PizerAnna ClericoAlessandra De GraziaAntonio PizzutiAnna MolesCarlo Dominici
Credits/Source: BMC Cancer 2014, 14:880