Distinct functional consequences of ECEL1/DINE missense mutations in the pathogenesis of congenital contracture...

Endothelin-converting enzyme-like 1 (ECEL1, also termed DINE in rodents), a membrane-bound metalloprotease, has been identified as a gene responsible for distal arthrogryposis (DA). ECEL1-mutated DA is generally characterized by ocular phenotypes in addition to the congenital limb contractures that are common to all DA subtypes. Until now, the consequences of the identified pathogenic mutations have remained incompletely understood because of a lack of detailed phenotypic analyses in relevant mouse models. In this study, we generated a new knock-in mouse strain that carries an ECEL1/DINE pathogenic G607S missense mutation, based on a previous study reporting atypical DA hindlimb phenotypes in two siblings with the mutation. We compared the morphological phenotypes of G607S knock-in mice with C760R knock-in mice that we previously established. Both C760R and G607S knock-in mouse embryos showed similar axonal arborization defects with normal trajectory patterns from the spinal cord to the target hindlimb muscles, as well as axon guidance defects of the abducens nerves. Intriguingly, distinct phenotypes in DINE protein localization and mRNA expression were identified in these knock-in mouse lines. For G607S, DINE mRNA and protein expression was decreased or almost absent in motor neurons. In the C760R mutant mice DINE was expressed and localized in the somata of motor neurons but not in axons. Our mutant mouse data suggest that ECEL1/DINE G607S and C760R mutations both lead to motor innervation defects as primary causes in ECEL1-mutated congenital contracture disorders. However, the functional consequences of the two mutations are distinct, with loss of axonal transport of ECEL1/DINE in C760R mutants and mRNA expression deficits in G607S mutants.