Masuma Akter

DYT1 dystonia is a debilitating neurological movement disorder, and it represents the most frequent and severe form of hereditary primary dystonia. There is currently no cure for this disease due to its unclear pathogenesis. In our previous study utilizing patient-specific motor neurons (MNs), we identified distinct cellular deficits associated with the disease, including a deformed nucleus, disrupted neurodevelopment, and compromised nucleocytoplasmic transport (NCT) functions. However, the precise molecular mechanisms underlying these cellular impairments have remained elusive. In this study, we revealed the genome-wide changes in gene expression in DYT1 MNs through transcriptomic analysis. We found that those dysregulated genes are intricately involved in neurodevelopment and various biological processes. Interestingly, we identified that the expression level of RANBP17, a RAN-binding protein crucial for NCT regulation, exhibited a significant reduction in DYT1 MNs. By manipulating RANBP17 expression, we further demonstrated that RANBP17 plays an important role in facilitating the nuclear transport of both protein and transcript cargos in induced human neurons. Excitingly, the overexpression of RANBP17 emerged as a substantial mitigating factor, effectively restoring impaired NCT activity and rescuing neurodevelopmental deficits observed in DYT1 MNs. These findings shed light on the intricate molecular underpinnings of impaired NCT in DYT1 neurons and provide novel insights into the pathophysiology of DYT1 dystonia, potentially leading to the development of innovative treatment strategies.

Generation of motor neurons (MNs) from human-induced pluripotent stem cells (hiPSCs) overcomes the limited access to human brain tissues and provides an unprecedent approach for modeling MN-related diseases. In this review, we discuss the recent progression in understanding the regulatory mechanisms of MN differentiation and their applications in the generation of MNs from hiPSCs, with a particular focus on two approaches: induction by small molecules and induction by lentiviral delivery of transcription factors. At each induction stage, different culture media and supplements, typical growth conditions and cellular morphology, and specific markers for validation of cell identity and quality control are specifically discussed. Both approaches can generate functional MNs. Currently, the major challenges in modeling neurological diseases using iPSC-derived neurons are: obtaining neurons with high purity and yield; long-term neuron culture to reach full maturation; and how to culture neurons more physiologically to maximize relevance to in vivo conditions.

DYT1 dystonia is a hereditary neurologic movement disorder characterized by uncontrollable muscle contractions. It is caused by a heterozygous mutation inTorsin A(TOR1A), a gene encoding a membrane-embedded ATPase. While animal models provide insights into disease mechanisms, significant species-dependent differences exist since animals with the identical heterozygous mutation fail to show pathology. Here, we model DYT1 by using human patient-specific cholinergic motor neurons (MNs) that are generated through either direct conversion of patients' skin fibroblasts or differentiation of induced pluripotent stem cells (iPSCs). These human MNs with the heterozygousTOR1Amutation show reduced neurite length and branches, markedly thickened nuclear lamina, disrupted nuclear morphology, and impaired nucleocytoplasmic transport (NCT) of mRNAs and proteins, whereas they lack the perinuclear “blebs” that are often observed in animal models. Furthermore, we uncover that the nuclear lamina protein LMNB1 is upregulated in DYT1 cells and exhibits abnormal subcellular distribution in a cholinergic MNs-specific manner. Such dysregulation of LMNB1 can be recapitulated by either ectopic expression of the mutantTOR1Agene or shRNA-mediated downregulation of endogenousTOR1Ain healthy control MNs. Interestingly, downregulation of LMNB1 can largely ameliorate all the cellular defects in DYT1 MNs. These results reveal the value of disease modeling with human patient-specific neurons and indicate that dysregulation of LMNB1, a crucial component of the nuclear lamina, may constitute a major molecular mechanism underlying DYT1 pathology.SIGNIFICANCE STATEMENTInaccessibility to patient neurons greatly impedes our understanding of the pathologic mechanisms for dystonia. In this study, we employ reprogrammed human patient-specific motor neurons (MNs) to model DYT1, the most severe hereditary form of dystonia. Our results reveal disease-dependent deficits in nuclear morphology and nucleocytoplasmic transport (NCT). Most importantly, we further identify LMNB1 dysregulation as a major contributor to these deficits, uncovering a new pathologic mechanism for DYT1 dystonia.