Complementation analysis in Gaucher disease using single cell microassay techniques. Evidence for a single “Gaucher gene”

Summary

Gaucher disease is a lysosomal storage disorder resulting from a deficiency of acid β-glucosidase. Several clinical forms have been described, including infantile, juvenile, and adult onset variants. We have examined complementation in infantile and adult forms of Gaucher disease by monitoring enzyme activity in multinucleate cells produced by fusing skin fibroblasts from different patients in the presence of polyethylene glycol. β-Glucosidase activity was monitored in lysates of individual multinucleate cells by a microassay method utilizing methylumbelliferyl-β-D-glucoside as the substrate (normal: 1.3±0.12x10^-13 mol/h/cell). The microassay was linear with time up to 4 h, for up to 20 mononucleate cells, and for individual multinucleate cells containing up to 12 nuclei. Complementation was examined in 11 fibroblasts strains fused in all pairwise combinations. In no instance was there any clear indication of complementation (at least 10–15% of normal activity to adequately account for experimental variation) although there was an indication of marginal increases in some fusions. On the other hand, the expected 50% activity was obtained in “heterozygous” fusions (normal/mutant) for both types of clinical variants. Our results are consistent with a single gene, presumably the structural gene encoding the enzyme, responsible for at least the infantile and adult variants, and confirm the autosomal recessive nature of the disorder.