Motor neuron disease due to neuropathy target esterase mutation: Enzyme analysis of fibroblasts from human subjects yields insights into pathogenesis

Abstract

Recently, we identified neuropathy target esterase (NTE) mutation as the cause of an autosomal recessive motor neuron disease (NTE-MND). Subsequently, we showed that NTE-MND mutations reduced specific activity (SA) and altered inhibitory kinetics of NTE catalytic domain constructs. Recent preliminary results showed that NTE is expressed in cultured human skin fibroblasts, and others have used mutant forms of neuronal proteins expressed in fibroblasts as biomarkers of neurogenetic diseases. Therefore, the present study was carried out to test the hypothesis that NTE in cultured skin fibroblasts from NTE-MND subjects also exhibit altered enzymological properties assessed by SA and IC 50 values of mipafox (MIP) and chlorpyrifos oxon (CPO). NTE SA was reduced to 65% of control (wild-type NTE from commercially obtained fibroblasts) in homozygous M1012V fibroblasts and 59–61% of control in compound heterozygous R890H/c2946_2947InsCAGC fibroblasts. MIP IC 50 values were unaffected by the NTE mutations, but the CPO IC 50 increased 4.5-fold in homozygous M1012V fibroblasts. Interestingly, markedly reduced NTE SAs (40–43% of control) were observed in fibroblasts from asymptomatic subjects heterozygous for NTE insertion c2946_2947InsCAGC. This insertion is predicted to produce truncated NTE missing the last 235 residues of its catalytic domain. These observations confirm that NTE-MND mutations reduce NTE SA in vitro. Moreover, to the extent observations made in cultured fibroblasts may be generalized to events in the nervous system, lack of correlation between reduced fibroblast NTE SA and the occurrence of NTE-MND in NTE insertion mutation heterozygotes indicates that reduction of NTE SA alone is insufficient to cause MND.