The Saccharomyces cerevisiae ICL2 gene encodes a mitochondrial 2-methylisocitrate lyase involved in propionyl-coenzyme A metabolism

The Saccharomyces cerevisiae ICL1 gene encodes isocitrate lyase, an essenti al enzyme for growth on ethanol and acetate. Previous studies have demonstr ated that the highly homologous ICL2 gene (YPR006c) is transcribed during t he growth of wild-type cells on ethanol. However, even when multiple copies are introduced, ICL2 cannot complement the growth defect of icl1 null muta nts. It has therefore been suggested that ICL2 encodes a nonsense mRNA or n onfunctional protein. In the methylcitrate cycle of propionyl-coenzyme A me tabolism, 2-methylisocitrate is converted to succinate and pyruvate, a reac tion similar to that catalyzed by isocitrate lyase. To investigate whether ICL2 encodes a specific 2-methylisocitrate lyase, isocitrate lyase and 2-me thylisocitrate lyase activities were assayed in cell extracts of wild-type S. cerevisiae and of isogenic icl1, icl2, and icl1 icl2 null mutants. Isoci trate lyase activity was absent in icl1 and icl1 icl2 null mutants, whereas in contrast, 2-methylisocitrate lyase activity was detected in the wild ty pe and single icl mutants but not in the icl1 icl2 mutant. This demonstrate d that ICL2 encodes a specific 2-methylisocitrate lyase and that the ICL1-e ncoded isocitrate lyase exhibits a low but significant activity with 2-meth ylisocitrate. Subcellular fractionation studies and experiments with an ICU -green fluorescent protein fusion demonstrated that the ICL2-encoded 2-meth ylisocitrate lyase is located in the mitochondrial matrix. Similar to that of ICL1, transcription of ICL2 is subject to glucose catabolite repression. In glucose-limited cultures, growth with threonine as a nitrogen source re sulted in a ca. threefold induction of ICL2 mRNA levels and of 2-methylisoc itrate lyase activity in cell extracts relative to cultures grown with ammo nia as the nitrogen source. This is consistent with an involvement of the 2 -methylcitrate cycle in threonine catabolism.