Pathomechanistic characterization of two exonic L1CAM variants located in trans in an obligate carrier of X-linked hydrocephalus

Mutations in the gene encoding the neural cell adhesion molecule L1CAM cause several neurological disorders collectively referred to as L1 syndrome. We report here a family case of X-linked hydrocephalus in which an obligate female carrier has two exonic L1CAM missense mutations in trans substituting amino acids in the first (p.W635C) or second (p.V768I) fibronectin-type III domains. We performed various biochemical and cell biological in vitro assays to evaluate the pathogenicity of these variants. Mutant L1-W635C protein accumulates in the endoplasmic reticulum (ER), is not transported into axons, and fails to promote L1CAM-mediated cell–cell adhesion as well as neurite growth. Immunoprecipitation experiments show that L1-W635C associates with the molecular ER chaperone calnexin and is modified by poly-ubiquitination. The mutant L1-V768I protein localizes at the cell surface, is not retained in the ER, and promotes neurite growth similar to wild-type L1CAM. However, the p.V768I mutation impairs L1CAM-mediated cell–cell adhesion albeit less severe than L1-W635C. These data indicate that p.W635C is a novel loss-of-function L1 syndrome mutation. The p.V768I mutation may represent a non-pathogenic variant or a variant associated with low penetrance. The poly-ubiquitination of L1-W635C and its association with the ER chaperone calnexin provide further insights into the molecular mechanisms underlying defective cell surface trafficking of L1CAM in L1 syndrome.