Abstract
Hereditary motor and sensory neuropathy with proximal dominant involvement (HMSN-P) is caused by a heterozygous mutation (P285L) in Tropomyosin-receptor kinase Fused Gene (TFG), histopathologically characterized by progressive spinal motor neuron loss with TFG cytosolic aggregates. Although the TFG protein, found as a type of fusion oncoprotein, is known to facilitate vesicle transport from endoplasmic reticulum (ER) to Golgi apparatus at ER exit site, it is unclear how mutant TFG causes motor neuron degeneration. Here we generated induced pluripotent stem cells (iPSCs) from HMSN-P patients, and differentiated the iPSCs into neural cells with spinal motor neurons (iPS-MNs). We found that HMSN-P patient iPS-MNs exhibited ubiquitin proteasome system (UPS) impairment, and HMSN-P patient iPS-MNs were vulnerable to UPS inhibitory stress. Gene correction of the mutation in TFG using the CRISPR-Cas9 system reverted the cellular phenotypes of HMSN-P patient iPS-MNs. Collectively, these results suggest that our cellular model with defects in cellular integrity including UPS impairments may lead to identification of pathomechanisms and a therapeutic target for HMSN-P.
Highlights
Hereditary motor and sensory neuropathy with proximal dominant involvement (HMSN-P) is an autosomaldominant disease first described in patients from the Okinawa Islands of Japan [1]
Generation of HMSN-P patient induced pluripotent stem cells (iPSCs) and spinal motor neurons (MNs) differentiation We generated iPSCs from healthy control subjects and HMSN-P patients with Tropomyosin-receptor kinase Fused Gene (TFG) P285L mutation (Table 1, Additional file 1: Figure S1A)
We analyzed mRNA expression levels of each of the iPSC clones using RNA-seq analysis, and confirmed that the significant differences were not observed among iPSCs generated from human dermal fibroblasts (HDFs) and iPSCs generated from peripheral blood mononuclear cells (PBMCs) (Additional file 1: Figure S1B), as shown previously [16]
Summary
Hereditary motor and sensory neuropathy with proximal dominant involvement (HMSN-P) is an autosomaldominant disease first described in patients from the Okinawa Islands of Japan [1]. The clinical manifestation of HMSN-P is characterized by late-onset progressive muscle weakness of proximal limbs with widespread fasciculations, shared with those of amyotrophic lateral sclerosis (ALS), and the patients eventually require artificial ventilation due to respiratory failure [2, 3]. A heterozygous mutation (P285L) in Tropomyosin-receptor kinase Fused Gene (TFG) was identified as the gene responsible for HMSN-P, and the pathological hallmark of HMSN-P is the formation of cytosolic inclusions of TFG protein in spinal MNs [5]. Homozygous TFG R106C mutation was identified in hereditary spastic paraplegia [9], and heterozygous TFG G269V mutation was found in Charcot-Marie-Tooth disease type 2 (CMT2) [10]
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