A Neuroligin-4 Missense Mutation Associated with Autism Impairs Neuroligin-4 Folding and Endoplasmic Reticulum Export

Abstract

Neuroligins (NLs) are postsynaptic cell-adhesion molecules essential for normal synapse function. Mutations in neuroligin-4 (NL4; gene symbol: NLGN4) have been reported in some patients with autism spectrum disorder (ASD) and other neurodevelopmental impairments. However, the low frequency of NL4 mutations, and the limited information about the affected patients and the functional consequences of their mutations, cast doubt on the causal role of NL4 mutations in these disorders. Here, we describe two brothers with classical ASD who carry a single amino acid substitution in NL4 (R87W). This substitution was absent from the brothers’ asymptomatic parents, suggesting that it arose in the maternal germline. R87 is conserved in all NL isoforms, and the R87W substitution is not observed in control individuals. At the protein level, the R87W-substitution impaired glycosylation processing of NL4 expressed in HEK293 and COS cells, destabilized NL4, caused NL4 retention in the endoplasmic reticulum in non-neuronal cells and neurons, and blocked NL4 transport to the cell-surface. As a result, the R87W-substitution inactivated the synapse-formation activity of NL4, and abolished the functional effect of NL4 on synapse strength. Viewed together, these observations suggest that a point mutation in NL4 can cause ASD by a loss-of-function mechanism.

Age-related expression of Neurexin1 and Neuroligin3 is correlated with presynaptic density in the cerebral cortex and hippocampus of male mice

“…Remarkably, mutations in Nlgn3/4 are involved in autism spectrum disorders (ASDs) which have been shown to destabilize the Nrxns-Nlgns complexes (Arac et al 2007;Chen et al 2008). In addition, a retention of Nlgns in the endoplasmic reticulum as well as a decrease in its affinity for Nrxns have been associated with some of those mutations (Comoletti et al 2004;Zhang et al 2009), suggesting that impaired trafficking and protein binding properties of Nrxns-Nlgns might be related to the disease.…”

Not all neuroligin 3 and 4X missense variants lead to significant functional inactivation

“…Vectors expressing a soluble IgG‐NX1β fusion protein (in a pCMV5 backbone) were provided by Professor Thomas C. Südhof (Zhang et al., 2009). …”

“…In vitro and in vivo experiments have indicated that the autism‐related neuroligin mutations may affect synapse maturation and function. (Bemben et al., 2015; Chih et al., 2004; Ey, Leblond, & Bourgeron, 2011; Jaramillo, Liu, Pettersen, Birnbaum, & Powell, 2014; Zhang et al., 2009). …”

The complex genetics in autism spectrum disorders

“…Multiple genetic changes in Nrxn [84][85][86][87][88][89][90][91][92][93][94] and Nlgn genes [92,93,[95][96][97][98][99][100][101][102][103][104][105][106][107][108][109][110] have been found in ASD patients. These changes include (i) point mutations, which cause frame shifts, small deletions, and missense mutations in both coding and promoter regions, (ii) distinct translocation events, and (iii) large-scale deletions of chromosomal DNA containing these gene loci.…”