Calcitriol, an active form of vitamin D, directly induces Rspo2 and prolongs survival of Chrne-deficient mouse, a model for congenital myasthenic syndrome. Mus musculus

Signal transduction at the the neuromuscular junction (NMJ) is compromised in a diverse array of diseases including congenital myasthenic syndromes (CMS). Germline mutations in CHRNE encoding the acetylcholine receptor (AChR) E subunit are the most common cause of CMS. An active form of vitamin D, calcitriol, binds to vitamin D receptor (VDR) and regulates gene expressions. We found that calcitriol enhanced MuSK phosphorylation, AChR clustering, and myotube twitching in in vitro NMJ comprised of C2C12 myotubes and NSC34 motor neurons. RNA-seq analysis of in vitro NMJ showed that calcitriol increased the expressions of Rspo2, Rapsn, and Dusp6. ChIP-seq of VDR revealed that VDR binds to a region approximately 15 kbp upstream to Rspo2. Biallelic deletion of the VDR-binding site of Rspo2 by CRISPR/Cas9 in C2C12 myoblasts/myotubes nullified the calcitriol-mediated induction of Rspo2 expression and MuSK phosphorylation. We generated a mouse line lacking Chrne by CRISPR/Cas9 as a model for CMS. Intraperitoneal administration of calcitriol markedly increased the number of AChR clusters, as well as the area, the intensity, and the number of synaptophysin-positive synaptic vesicles, in Chrne KO mice. In addition, calcitriol ameliorated motor deficits and prolonged survival of Chrne-deficient mice. In the skeletal muscle, calcitriol increased the gene expressions of Rspo2, Rapsn, and Dus6. We propose that calcitriol is a potential therapeutic agent for CMS and other diseases with defective neuromuscular signal transmission.