Novel insights into the molecular mechanisms of LGMDD2: role of TNPO3 in experimental cell and zebrafish models.

Transportin 3 (TNPO3) is a nuclear carrier for serine/arginine-rich proteins which are essential for mRNA splicing and metabolism. Mutations in the TNPO3 gene result in a protein with an extended C-terminal domain, leading to the onset of LGMDD2, a rare form of limb girdle muscular dystrophy. To investigate the role of TNPO3 in skeletal muscle and the pathogenic mechanism underlying LGMDD2, we develped both in vitro and in vivo models of the disease. The in vitro model was established using the C2C12 cell line, transfected with plasmids encoding either the wild-type (WT) or mutated (MUT) human TNPO3 (hTNPO3). For the in vivo model, we microinjected zebrafish (Danio rerio) embryos with mRNAs encoding WT or MUT hTNPO3. We analyzed the expression patterns of myogenesis-related genes, muscle-specific genes, myomiRNA and genes strictly related to the disease. These analyses were complemented by protein expression studies, morphological assessments in both models, and behavioral assays to validate the LGMDD2 zebrafish model. Our results demonstrate a key role for TNPO3 in regulating myogenesis in both models and reveal that the TNPO3 mutation disrupts normal myogenic commitment, supporting its contribution to LGMDD2 pathogenesis. Overall, this study represents a significant advance in understanding the role of TNPO3 in skeletal muscle biology and the molecular basis of LGMDD2.