Evolutionary engineering and molecular characterization of a caffeine-resistant Saccharomyces cerevisiae strain
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Caffeine is a naturally occurring alkaloid, where its major consumption occurs with beverages such as coffee, soft drinks and tea. Despite a variety of reports on the effects of caffeine on diverse organisms including yeast, the complex molecular basis of caffeine resistance and response has yet to be understood. In this study, a caffeine-hyperresistant and genetically stable Saccharomyces cerevisiae mutant was obtained for the first time by evolutionary engineering, using batch selection in the presence of gradually increased caffeine stress levels and without any mutagenesis of the initial population prior to selection. The selected mutant could resist up to 50 mM caffeine, a level, to our knowledge, that has not been reported for S. cerevisiae so far. The mutant was also resistant to the cell wall-damaging agent lyticase, and it showed cross-resistance against various compounds such as rapamycin, antimycin, coniferyl aldehyde and cycloheximide. Comparative transcriptomic analysis results revealed that the genes involved in the energy conservation and production pathways, and pleiotropic drug resistance were overexpressed. Whole genome re-sequencing identified single nucleotide polymorphisms in only three genes of the caffeine-hyperresistant mutant; PDR1, PDR5 and RIM8, which may play a potential role in caffeine-hyperresistance.
KeywordsAdaptive laboratory evolution Caffeine Evolutionary engineering Pleiotropic drug resistance (PDR) Saccharomyces cerevisiae Stress resistance
We thank Levent Üge for technical assistance with HPLC analyses, and our former and present students Ogün Morkoç, Gizem Karabıyık, Diler Kaan Atmaca, and İsmail Can Karaoğlu for their help with the physiological experiments. We also thank Cihan Erdinç Gülsev and Nazlı Kocaefe for technical assistance with the whole genome re-sequencing experiments, Burcu Hacısalihoğlu for fruitful discussions and experimental assistance, Prof. Dr. Oğuz Öztürk for providing propolis and Prof. Dr. Nevin Gül Karagüler for her helpful comments regarding our SNP data.
The authors received no financial support for the research presented in this article.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
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