Evolution of a glucose-regulated ADH gene in the genus Saccharomyces

To determine when a glucose-repressed alcohol dehydrogenase isozyme and its regulatory gene, ADR1, arose during evolution, we surveyed species of the genus Saccharomyces for glucose-repressed ADH isozymes and for ADR1 homolog ues. Glucose repressed ADH isozymes were present in all species of Saccharo myces sensu strictu and also in Saccharomyces kluyveri, the most distant me mber of the Saccharomyces clade. We cloned and characterized ADH promoters from S. bayanus, S. douglasii, and S. kluyveri. The ADH promoters from S. b ayanus and S. douglasii had conserved sequences, including upstream regulat ory elements, and an extended polydA tract. The expression of a reporter ge ne driven by the S. bayanus promoter was glucose-repressed and dependent on the major activator of transcription, ADR1, when it was introduced into S. cerevisiae. One S. kluyveri promoter was also glucose-repressed and ADR1-d ependent in S. cerevisiae. The other S. kluyveri ADH promoter was expressed constitutively and was ADR1-independent. Although showing little sequence conservation with the S. cerevisiae ADH2 promoter, the glucose-repressed S. kluyveri promoter contains numerous potential binding sites for Adr1. The glucose-repressed ADH from S. kluyveri is a mitochondrial isozyme most clos ely related to S. cerevisiae ADHIII. ADR1 homologues from S. douglasii and S. paradoxus contain a trinucleolide repeat encoding polyAsn that is lackin g in S. cerevisiae and S. bayanus. No ADR1 homologue could be detected in S . kluyveri, suggesting that the potential for Adr1 regulation may have aris en first, before ADR1 evolved. (C) 2000 Elsevier Science B.V. All rights re served.