In Kluyveromyces lactis a Pair of Paralogous Isozymes Catalyze the First Committed Step of Leucine Biosynthesis in Either the Mitochondria or the Cytosol

Divergence of paralogous pairs, resulting from gene duplication, plays an important role in the evolution of specialized or novel gene functions. Analysis of selected duplicated pairs has elucidated some of the mechanisms underlying the functional diversification ofSaccharomyces cerevisiae(S.cerevisiae) paralogous genes. Similar studies of the orthologous pairs extant in pre-whole genome duplication yeast species, such asKluyveromyces lactis(K. lactis) remain to be addressed. The genome ofK. lactis, an aerobic yeast, includes gene pairs generated by sporadic duplications. The genome of this organism comprises theKlLEU4andKlLEU4BISparalogous pair, annotated as putative alpha-isopropylmalate synthases (alpha-IPMSs), considered to be the orthologs of theS. cerevisiae ScLEU4/ScLEU9paralogous genes. The enzymes encoded by the latter two genes are mitochondrially located, differing in their sensitivity to leucine allosteric inhibition resulting inScLeu4-ScLeu4 andScLeu4-ScLeu9 sensitive dimers andScLeu9-ScLeu9 relatively resistant homodimers. Previous work has shown that, in aScleu4 Delta mutant,ScLEU9expression is increased and assembly ofScLeu9-ScLeu9 leucine resistant homodimers results in loss of feedback regulation of leucine biosynthesis, leading to leucine accumulation and decreased growth rate. Here we report that: (i)K. lactisharbors a sporadic gene duplication, comprising theKlLEU4, syntenic withS. cerevisiae ScLEU4andScLEU9, and the non-syntenicKlLEU4BIS, arising from a pre-WGD event. (ii) That both,KlLEU4andKlLEU4BISencode leucine sensitive alpha-IPMSs isozymes, located in the mitochondria (KlLeu4) and the cytosol (KlLeu4BIS), respectively. (iii) That both,KlLEU4orKlLEU4BIScomplement theScleu4 Delta Scleu9 Delta leucine auxotrophic phenotype and revert the enhancedScLEU9transcription observed in aScleu4 Delta ScLEU9mutant. TheScleu4 Delta ScLEU9growth mutant phenotype is only fully complemented when transformed with the syntenicKlLEU4mitochondrial isoform.KlLEU4andKlLEU4BISunderwent a different diversification pathways than that leading toScLEU4/ScLEU9.KlLEU4could be considered as the functional ortholog ofScLEU4, since its encoded isozyme can complement both theScleu4 Delta Scleu9 Delta leucine auxotrophy and theScleu4 Delta ScLEU9complex phenotype.