Genetic complementation in heterokaryons of human fibroblasts defective in cobalamin metabolism.
Inherited methylmalonicacidemia due to deficiency of methylmalonyl-CoA mutase (methylmalonyl-CoA CoA-carbonylmutase; EC 5.4.99.2) activity results from at least three classes of biochemically distinct defects affecting cobalamin (Cbl: vitamin B12) metabolism (cbl A, cbl B, and cbl C mutants) and a fourth class producing a defective mutase apoenzyme. We have obtained genetic evidence in support of this biochemical heterogeneity, using heterokaryons prepared by Sendai-virus-mediated cell fusion. Nine fibroblast lines from patients with defective Cbl metabolism (4 cbl A, 3 cbl B, and 2 cbl C), two from patients with defective mutase apoenzyme, and two from controls were fused in pairwise combinations and tested for functional mutase holoenzyme using a radioautographic procedure which detects [14C]propionate incorporation into trichloroacetic-acid-precipitable material in fibroblast monolayers in situ. Each of the mutants incorporates negligible radioactivity compared to control cells. Activity is also negligible when different mutants are mixed without virus or when homokaryons are produced by self-fusion. Heterokaryons produced by fusing members of each of the four mutant classes with representatives of any other class recover the ability to incorporate [14C]propionate to levels comparable to those of control cells. However, heterokaryons produced between members of the same class fail to complement in all cases. We conclude that the mutants with defective Cbl metabolism (cbl A, cbl B, cbl C) comprise three complementation groups, that a fourth group corresponds to mutase apoenzyme deficiency, and that all four classes of mutations are recessively inherited.
D000592:Amino Acid Metabolism, Inborn Errors / Q000235:genetics*; D002459:Cell Fusion; D002478:Cells, Cultured; D005347:Fibroblasts / Q000378:metabolism* / Q000648:ultrastructure; D005796:Genes; D005816:Genetic Complementation Test; D006579:Heterozygote; D008764:Methylmalonic Acid / Q000378:metabolism; D009154:Mutation; D011422:Propionates / Q000378:metabolism; D012867:Skin / Q000378:metabolism*; D014805:Vitamin B 12 / Q000378:metabolism*
D011422:Propionates; D008764:Methylmalonic Acid; D014805:Vitamin B 12