Laboratory evolution of new lactate transporter genes in a jen1Δ mutant of Saccharomyces cerevisiae and their identification as ADY2 alleles by whole-genome resequencing and transcriptome analysis.

Autor: Kok, Stefan, Nijkamp, Jurgen F., Oud, Bart, Roque, Filipa C., Ridder, Dick, Daran, Jean-Marc, Pronk, Jack T., Maris, Antonius J. A.
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Zdroj: FEMS Yeast Research; May2012, Vol. 12 Issue 3, p359-374, 16p
Abstrakt: Laboratory evolution is a powerful approach in applied and fundamental yeast research, but complete elucidation of the molecular basis of evolved phenotypes remains a challenge. In this study, DNA microarray-based transcriptome analysis and whole-genome resequencing were used to investigate evolution of novel lactate transporters in Saccharomyces cerevisiae that can replace Jen1p, the only documented S. cerevisiae lactate transporter. To this end, a jen1Δ mutant was evolved for growth on lactate in serial batch cultures. Two independent evolution experiments yielded growth on lactate as sole carbon source (0.14 and 0.18 h−1, respectively). Transcriptome analysis did not provide leads, but whole-genome resequencing showed different single-nucleotide changes ( C755 G/ Leu219 Val and C655 G/ Ala252 Gly) in the acetate transporter gene ADY2. Introduction of these ADY2 alleles in a jen1Δ ady2Δ strain enabled growth on lactate (0.14 h−1 for Ady2pLeu219Val and 0.12 h−1 for Ady2pAla252Gly), demonstrating that these alleles of ADY2 encode efficient lactate transporters. Depth of coverage of DNA sequencing, combined with karyotyping, gene deletions and diagnostic PCR, showed that an isochromosome III ( c. 475 kb) with two additional copies of ADY2C755G had been formed via crossover between retrotransposons YCLWΔ15 and YCRCΔ6. The isochromosome formation shows how even short periods of selective pressure can cause substantial karyotype changes. [ABSTRACT FROM AUTHOR]
Databáze: Complementary Index
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