The dimorphic yeast Yarrowia lipolytica is more resistant to high copper concentrations than Saccharomyces cerevisiae. This differential tolerance to copper ions has been observed in several strains arising from non-related isolates. To investigate the molecular basis of this resistance, we obtained several copper-sensitive mutants. By complementation of one of them, we isolated the YlCRF1 gene encoding for a copper-binding transcription factor of 411 amino acids homologous to ScAce1p, CgAmt1p, and ScMac1p. Naturally occurring copper-sensitive strains lack the CRF1 allele. The YlCRF1 transcript is not induced by the addition of copper to the medium. Gene disruption demonstrated that YlCRF1 is responsible for a 4- to 5-fold increase in Y. lipolytica copper tolerance. We further show that strain Deltacrf1 is more sensitive to cadmium but not to other metals. The role of YlCrf1p as a copper-sensitive transcription factor is supported by the finding that the protein is immunolocalized in the nucleus during growth in copper-supplemented but not in copper-free medium. However, in contrast to the S. cerevisiae strain mutated in the metallothionein transcription activator ACE1, Y. lipolytica strain Deltacrf1 is still able to increase metallothionein (MTP) mRNA levels in response to copper addition. CRF1 deletion does not affect superoxide dismutase (SOD) activity either. Our data suggest the existence of one or more different target genes for Crf1p, other than MTP or SOD1, and support its role as a novel copper-responsive transcription factor involved in metal detoxification.

• PMID: 12114501
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Comparative Study | Journal Article | Research Support, Non-U.S. Gov't

Authors

García S, Prado M, Dégano R, Domínguez A

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