Postnatal knockdown of dok‐7 gene expression in mice causes structural defects in neuromuscular synapses and myasthenic pathology

Abstract

The neuromuscular junction (NMJ) is a synapse between a motor neuron and skeletal muscle and is required for muscle contraction. The formation and maintenance of NMJs are governed by the muscle-specific receptor tyrosine kinase MuSK. We previously showed that the muscle cytoplasmic protein Dok-7 is an essential activator of MuSK. Indeed, mice lacking either Dok-7 or MuSK form no NMJs, and defects in the human DOK7 gene underlie a congenital myasthenic syndrome (an NMJ disorder). However, it remains unproven whether Dok-7 is required for the postnatal maintenance of NMJs. In this study, we generated recombinant adeno-associated virus (AAV) vectors encoding short hairpin RNAs targeting the mouse dok-7 gene (AAV-shD7). Systemic administration of AAV-shD7 into 2-week-old mice down-regulated dok-7 expression in muscle and induced myasthenic symptoms including reduction in body weight and motor function. Moreover, AAV-shD7 treatment suppressed MuSK-dependent gene expression of NMJ components and reduced the size of NMJs. These results demonstrate that correct, physiological levels of dok-7 expression are required for the postnatal maintenance of NMJs.

The beta-adrenergic agonist salbutamol modulates neuromuscular junction formation in zebrafish models of human myasthenic syndromes

“…The relatively short exposure duration of the zebrafish to salbutamol and forskolin may further explain why only a partial rescue of the phenotype was observed in both models. In addition, patients with CMS due to DOK7 mutations frequently present after infancy or in early adulthood ( 38 ), and postnatal knockdown of Dok-7 gene expression has been shown to cause CMS in mice ( 39 ), indicating a further role for Dok-7-mediated activation of MuSK in postnatal NMJ maintenance. Therefore, it is possible that the improvement seen in β-agonist-treated CMS patients is due to an additional role they play in NMJ homeostasis.…”

NMJ maintenance and repair in aging

“…These examples highlight the interdependence between all three cellular components of the NMJ, and roles of synapse-associated molecules in the maintenance and repair of adult NMJs. In this regard, studies over the last few years have shown that synaptic molecules agrin, Lrp4, the muscle-specific kinase (MuSK) receptor, and Dok7 remain essential at adult NMJs [61–64]. For example, deletion of agrin in a subset of adult motor neurons causes the disintegration of the postsynapse and degeneration of motor axons [63].…”

“…For example, deletion of agrin in a subset of adult motor neurons causes the disintegration of the postsynapse and degeneration of motor axons [63]. Likewise, adult NMJs degenerate in the absence of LRP4, MuSK, or Dok7 [61,62,64]. Thus, molecules required for the formation of the NMJ continue to play integral functions in adulthood.…”

The Mouse Levator Auris Longus Muscle: An Amenable Model System to Study the Role of Postsynaptic Proteins to the Maintenance and Regeneration of the Neuromuscular Synapse

“…Following a facial nerve crush injury protocol, we observed axonal degeneration and muscle fiber reinnervation similar to that described in the hind limb Tibialis anterior muscle after sciatic nerve crush injury (Magill et al, 2007 ). In the context of NMJ regeneration, several muscle proteins that play essential roles in the embryonic assembly have demonstrated a potentially key role on helping NMJ maintenance, such as LRP4 (Barik et al, 2014 ), Dok-7 (Eguchi et al, 2016 ), and rapsyn (Kong et al, 2004 ; Martínez-Martínez et al, 2009 ). Indeed, overexpression of a low copy number of MuSK partially rescues the NMJ degeneration that takes place at pre-symptomatic stages of Amyotrophic Lateral Sclerosis animal models (Pérez-Garcia and Burden, 2012 ).…”