KlADH3 and KlADH4 are Kluyveromyces lactis genes encoding the two mitochondrial alcohol dehydrogenase activities located within mitochondria. In this yeast, ethanol induces the transcription of KlADH4 and, conversely, represses that of KlADH3. In this study, we describe the effects of the aar900 mutation on such regulation. This mutation, firstly isolated in a strain devoid of alcohol dehydrogenase genes except KlADH4, conferred to cells resistance to allyl alcohol because of the absence of the KlAdh4p activity. When the mutation was transferred by crosses to an isogenic strain containing all the alcohol dehydrogenase genes, we found that the KlADH3 gene was highly expressed even in the presence of ethanol. In addition, we observed that the absence of KlAdh4p resulted from a post-transcriptional control in that KlADH4 was transcriptionally induced by ethanol. We also found that KlPDC1, another ethanol-repressible gene, was not transcribed in the mutant in the presence of this carbon source, indicating that the escape of KlADH3 from ethanol repression was a peculiar feature of this gene. Genetic analysis showed a Mendelian segregation of the mutation that was mapped in a region of chromosome III close to the ade1 locus. Interestingly, the aar900 mutants had a pleiotropic phenotype and showed increased resistance to monovalent cations and benomyl, suggesting that the mutation could also affect genes other than the alcohol dehydrogenase ones. Strains carrying the aar900 mutation could represent useful tools to unravel the peculiar regulation of KlADH3 and KlADH4 by ethanol.