Structure and regulatory expression of a single copy alternative oxidase gene from the yeast Pichia anomala.

Abstract

To investigate the regulatory mechanism of alternative oxidase gene expression, genomic DNA was cloned from the yeast Pichia anomala. Genomic Southern blot analysis suggested that a single copy nuclear gene encoded an alternative oxidase in the yeast. The nucleotide sequence showed an uninterrupted coding region for the alternative oxidase protein. In the upstream region from the transcription initiation site found by primer extension analysis, CCAAT, TATAA, and UAS2-like elements were detected. The UAS2 is the element involved in transcriptional regulation by carbon source and the target site for the factor, HAP2/3/4/5 protein complex, in Saccharomyces cerevisiae. By a gel mobility shift assay, a specific retardation band was detected when a protein extract from cells grown on an inducing carbon source was incubated with a UAS2-containing probe. These results suggest that carbon source regulation of alternative oxidase gene expression is mediated by the UAS2-like element and a HAP-like factor in P. anomala.