Transcriptomic and morphological alterations in hTNPO3 MUT-microinjected Zebrafish embryos modelling Limb Girdle Muscular Dystrophy D2 in vivo

Transportin-3, encoded by the TNPO3 gene, is physiologically involved in the translocation to the nucleus of many different proteins, such as SR proteins, which are associated with RNA metabolism. Mutations in the termination codon of the TNPO3 gene have been proved to cause Limb Girdle Muscular Dystrophy D2 (LGMD D2). More specifically, a single nucleotide deletion leads to the production of a protein with a 15 amino acid extension of its C-terminal domain. This project aimed to investigate the cause of LGMD D2, utilizing an in vivo approach, focusing on the role of TNPO3 on muscle development and to identify any potential molecular pathways linked to the disorder. This procedure involved the microinjection of mRNAs encoding the mutated form of human TNPO3 in Zebrafish embryos, to follow its effects on the myogenic processes during development. The subsequent analysis revealed changes in the gene expression profiles of Myogenic Regulatory Factors (MRFs) and muscle-specific proteins. The transcriptomic alterations are reflected on a phenotypical level by an aberrant organization of muscle fibres, as demonstrated by immunofluorescence staining. The methods we employed enabled us to gain an initial understanding of the role of Transportin-3 in muscle development and the pathogenic mechanism of its mutated form underlying LGMD D2, which has yet to be determined.