Distal hereditary motor neuropathies (dHMNs) are a group of inherited diseases involving the progressive, length-dependent axonal degeneration of the lower motor neurons. There are currently 29 reported causative genes and 4 disease loci implicated in dHMN. Despite the high genetic heterogeneity, mutations in the known genes account for less than 20% of dHMN cases with the mutations identified predominantly being point mutations or indels. We have expanded the spectrum of dHMN mutations with the identification of a 1.35 Mb complex structural variation (SV) causing a form of autosomal dominant dHMN (DHMN1 OMIM %182906). Given the complex nature of SV mutations and the importance of studying pathogenic mechanisms in a neuronal setting, we generated a patient-derived DHMN1 motor neuron model harbouring the 1.35 Mb complex insertion. The DHMN1 complex insertion creates a duplicated copy of the first 10 exons of the ubiquitin-protein E3 ligase gene (UBE3C) and forms a novel gene-intergenic fusion sense transcript by incorporating a terminal pseudo-exon from intergenic sequence within the DHMN1 locus. The UBE3C intergenic fusion (UBE3C-IF) transcript does not undergo nonsense-mediated decay and results in a significant reduction of wild type full length UBE3C (UBE3C-WT) protein levels in DHMN1 iPSC-derived motor neurons. An engineered transgenic C. elegans model expressing the UBE3C-IF transcript in GABA-ergic motor neurons shows neuronal synaptic transmission deficits. Furthermore, the transgenic animals are susceptible to heat stress which may implicate defective protein homeostasis underlying DHMN1 pathogenesis. Identification of the novel UBE3C-IF gene-intergenic fusion transcript in motor neurons highlights a potential new disease mechanism underlying axonal and motor neuron degeneration. These complementary models serve as a powerful paradigm for studying the DHMN1 complex SV and an invaluable tool for defining therapeutic targets for DHMN1.

Background Heterozygous KMT2B variants are a common cause of dystonia. A novel synonymous KMT2B variant, c.5073C>T (p.Gly1691=) was identified in an individual with childhood‐onset progressive dystonia. Methods The splicing impact of c.5073C>T was assessed using an in vitro exon‐trapping assay. The genomic region of KMT2B exons 23–26 was cloned into the pSpliceExpress plasmid between exon 2 and 3 of the rat Ins2 gene. The c.5073C>T variant was then introduced through site‐directed mutagenesis. The KMT2B wild‐type and c.5073C>T plasmids were transfected separately into HeLa cells and RNA was extracted 48 hours after transfection. The RNA was reverse transcribed to produce cDNA, which was PCR amplified using primers annealing to the flanking rat Ins2 sequences. Results Sanger sequencing of the PCR products revealed that c.5073C>T caused a novel splice donor site and therefore a 5‐bp deletion of KMT2B exon 23 in mature mRNA, leading to a coding frameshift and premature stop codon (p.Lys1692AsnfsTer7). Conclusion To our knowledge, this is the first report of a KMT2B synonymous variant associated with dystonia. Reassessment of synonymous variants may increase diagnostic yield for inherited disorders including monogenic dystonia. This is of clinical importance, given the generally favourable response to deep brain stimulation for KMT2B ‐related dystonia.

EGR2 (early growth response 2) is a crucial transcription factor for the myelination of the peripheral nervous system. Mutations in EGR2 are reported to cause a heterogenous spectrum of peripheral neuropathy with wide variation in both severity and age of onset, including demyelinating and axonal forms of Charcot-Marie Tooth (CMT) neuropathy, Dejerine-Sottas neuropathy (DSN/CMT3), and congenital hypomyelinating neuropathy (CHN/CMT4E). Here we report a sporadic de novo EGR2 variant, c.1232A > G (NM_000399.5), causing a missense p.Asp411Gly substitution and discovered through whole-exome sequencing (WES) of the proband. The resultant phenotype is severe demyelinating DSN with onset at two years of age, confirmed through nerve biopsy and electrophysiological examination. In silico analyses showed that the Asp411 residue is evolutionarily conserved, and the p.Asp411Gly variant was predicted to be deleterious by multiple in silico analyses. A luciferase-based reporter assay confirmed the reduced ability of p.Asp411Gly EGR2 to activate a PMP22 (peripheral myelin protein 22) enhancer element compared to wild-type EGR2. This study adds further support to the heterogeneity of EGR2-related peripheral neuropathies and provides strong functional evidence for the pathogenicity of the p.Asp411Gly EGR2 variant. Charcot-Marie-Tooth (CMT) neuropathy is group of degenerative motor and sensory peripheral neuropathies which are clinically and genetically heterogenous. Pathogenic variants in over 90 genes cause CMT, and whole-exome sequencing (WES) is now an effective tool for screening known causative genes in unsolved CMT families. Pathogenic variants in the EGR2 gene (early growth response 2) cause a broad spectrum of peripheral neuropathy phenotypes. This includes two forms of severe early-onset peripheral neuropathy, Dejerine-Sottas neuropathy (DSN/CMT3) 1-3 and congenital hypomyelinating neuropathy (CHN/CMT4E) 4,5 , as well as adult-onset demyelinating CMT1D with mild-moderate symptoms 3-12 and variable-onset axonal CMT with varied symptom severity 13,14. EGR2 encodes a C 2 H 2-type zinc-finger transcription factor that regulates the expression of genes involved in the formation and maintenance of myelin, including GJB1 (gap junction beta 1), MPZ (myelin protein zero), MBP (myelin basic protein), MAG (myelin associated glycoprotein), PRX (periaxin), and PMP22 (peripheral myelin protein 22) 4,15-18. Approximately half of the reported disease-associated EGR2 variants are de novo and are associated with a severe phenotype 13. Here, we report a sporadic de novo EGR2 variant in the third zinc-finger domain, c.1232A > G p.Asp411Gly, discovered through WES candidate gene screening. This variant manifested as a severe DSN phenotype with early onset at two years of age.