Data di Pubblicazione:
2015
Citazione:
RUOLO DEI MICRORNA NEL DIFFERENZIAMENTO MIOGENICO NORMALE E PATOLOGICO / F. Roma ; tutor: E. Ginelli ; coordinator: M. Locati. DIPARTIMENTO DI BIOTECNOLOGIE MEDICHE E MEDICINA TRASLAZIONALE, 2015 Jan 12. 27. ciclo, Anno Accademico 2014. [10.13130/roma-francesca_phd2015-01-12].
Abstract:
riassunto Facioscapulohumeral muscular dystrophy (FSHD) is an autosomal dominant disease, is the third most common myopathy, with an incidence of 1 in 15.000 in the general population. Clinical symptoms appear during the second decade of age and are characterized by progressive muscle weakness and atrophy, initially of the facial, scapular and humeral muscles, which show marked asymmetry and later involving the abdominal muscles and the musculature of the lower limbs. This disease is generally associated with contraction of D4Z4 repeat array (1 to 10 repeats), localized on chromosome 4 (4q35 region). D4Z4 is a macrosatellite typically composed of 10 to 100 of 3.3 kb units repeated in tandem. Every unit has a complex structure, with GC-rich sequences and an open reading frame (ORF), containing two homeobox sequences. Upstream of the macrosatellite are located several genes (FRG1, FRG2, ANT1) normally not expressed, or expressed at very low levels. The whole region is indeed strongly methylated, with an overall heterochromatic state. The contraction, together or in addition to molecular mechanisms not yet totally clarified, leads to 4q35 hypomethylation, causing gene overexpression of DUX4 and another genes in cis and trans. Infact, the FSHD cells are characterized by an extensive gene expression dysregulation mainly affecting the myogenesis. In this context, miRNAs may play an important role, are powerful gene expression modulators. Moreover, there are muscle-specific microRNAs (myomiRNAs), actively regulating several myogenic factors, as MEF2, MyoD, myogenin and SRF.
This thesis proposes to relate differentiation processes of both healthy and FSHD myoblasts, investigating the expression prolifes, by RNA-Seq, of miRNA . The purpose is indeed to clarify possible expression differences, the derived miRNA prolifing could represent a novel molecular signature for FSHD that includes diagnostic biomarkers and possibly therapeutic targets. MicroRNAs with a significantly different expression profile between FSHD and controls have been submitted to qRT-PCR, in order to validate the sequencing results. The subsequent investigation by bioinformatics prediction tools on the targets of these small ncRNA, allowed to define molecular pathways that could be altered in FSHD. In short, control myogenesis showed the modulation of 38 miRNAs, the majority of which (34 out 38) were up-regulated, including myomiRs (miR-1, -133a, -133b and -206). Approximately one third of the modulated miRNAs were not previously reported to be involved in muscle differentiation, and interestingly some of these ( i.e. miR-874, -1290,-95 and -146a) were previously shown to regulate cell proliferation and differentiantion. FSHD myogenesis evidenced a reduced number of modulated miRNAs than healthy muscle cells. The two processes shared nine miRNAs, including myomiRs, although with FC values lower in FSHD than in control cells. In addition, FSHD cells showed the modulation of six miRNAs ( miR-1268, -1268b, -1908, -4258, -4508 and -4516) not evidenced in control cells and that therefore could be considered FSHD-specific, likewise three novel miRNAs that seem to be specifically expressed in FSHD myotubes. These data further clarify the impact of miRNA regulation during control myogenesis and strongly suggest that a complex dysregulation of miRNA expression characterizes FSHD, impairing two important features of myogenesis: cell cycle and muscle development. However, the full range of molecular alterations at the basis of FSHD is not yet fully deciphered and the results can be a starting point for future research focused towards this goal. Among the various approaches to the attempt to understand the pathophysiological mechanism of dystrophy you can include the derivation
Tipologia IRIS:
Tesi di dottorato
Elenco autori:
F. Roma
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