[P3-111] Identification of Genes Involving Neuromuscular Junction Impairment in Taiwanese Spinal Muscular Atrophy Mouse Model by RNA-Seq
[Introduction]Spinal muscular atrophy (SMA), caused by deletion/mutations of survival motor neurons 1 (SMN1) gene, is characterized by progressive motor neuron (MN) degeneration in spinal cord. It has been hypothesized that neuromuscular junction (NMJ) defect might be involved in SMA pathogenesis. However, the molecular basis of NMJ defects in SMA pathogenesis has remained unresolved.
[Methods and Results] RNA-Sequencing (RNA-Seq) was performed on spinal cord samples of 3 severe Taiwanese SMA mice (Smn-/-; hSMN2tg/0) and 3 wild-type controls at postnatal day (P9), respectively. Subsequently, the reverse transcription-PCR(RT-PCR) was applied to validate the results of RNA-Seq. We identified related downstream effectors whether were affected by the dysregulated upstream, showing molecular events that could explain key aspects of NMJ pathology in SMA. RNA-Seq revealed 800 potential genes expressing differentially between SMA and control mice, including 642 down-regulated and 158 up-regulated genes. Six NMJ-associated candidate genes were validated by RT-PCR at P2, P5 and P9, respectively. The results indicated a strongest effect of integrin beta 1 (Itgb1). However, suppression of the downstream target of Itgb1, Abelson-related gene protein (Arg), did not activate RhoA/ROCK and cofilin. Otherwise, downregulation of PTK2 (protein tyrosine kinase 2) and FARP2 (FERM, ARH/RhoGEF and Pleckstrin domain protein 2) inactivated RhoA/ROCK pathway.
[Conclusions] In this study, we found 800 significantly differential genes by RNA-Seq. We further identify that the focal adhesion pathway might involve in SMA pathogenesis, and Itgb1, a crucial NMJ-associated gene, may be a potential therapeutic target of SMA.
[Methods and Results] RNA-Sequencing (RNA-Seq) was performed on spinal cord samples of 3 severe Taiwanese SMA mice (Smn-/-; hSMN2tg/0) and 3 wild-type controls at postnatal day (P9), respectively. Subsequently, the reverse transcription-PCR(RT-PCR) was applied to validate the results of RNA-Seq. We identified related downstream effectors whether were affected by the dysregulated upstream, showing molecular events that could explain key aspects of NMJ pathology in SMA. RNA-Seq revealed 800 potential genes expressing differentially between SMA and control mice, including 642 down-regulated and 158 up-regulated genes. Six NMJ-associated candidate genes were validated by RT-PCR at P2, P5 and P9, respectively. The results indicated a strongest effect of integrin beta 1 (Itgb1). However, suppression of the downstream target of Itgb1, Abelson-related gene protein (Arg), did not activate RhoA/ROCK and cofilin. Otherwise, downregulation of PTK2 (protein tyrosine kinase 2) and FARP2 (FERM, ARH/RhoGEF and Pleckstrin domain protein 2) inactivated RhoA/ROCK pathway.
[Conclusions] In this study, we found 800 significantly differential genes by RNA-Seq. We further identify that the focal adhesion pathway might involve in SMA pathogenesis, and Itgb1, a crucial NMJ-associated gene, may be a potential therapeutic target of SMA.