A novel arabinose-inducible promoter in the carotenogenic yeast Xanthophyllomyces dendrorhous
TL;DR
It is believed that the LAD promoter is a promising tool for fine-tuning of protein expression in X. dendrorhous, a basidiomycetous yeast capable of synthesizing astaxanthin, a powerful antioxidant carotenoid pigment.
Abstract
Xanthophyllomyces dendrorhous is a basidiomycetous yeast capable of synthesizing astaxanthin, a powerful antioxidant carotenoid pigment. Despite appreciable research efforts, molecular tools for the genetic manipulation of this yeast remain limited. Inducible promoters are essential tools to control gene expression in response to external stimuli, such as molecules or stress signals. This regulation enables precise control of gene expression in synthetic biological systems, minimizing unwanted effects caused by constant gene expression. In the present work inducible promoters from the L-arabinitol dehydrogenase ( LAD ), isocitrate lyase ( ICL ), alpha glucosidase ( AG ), glycerol kinase ( GUT1 ), and xylitol dehydrogenase ( XDH ) genes were screened and evaluated. The promoter sequences were cloned into a plasmid containing the encoding sequence for Enhanced Yellow Fluorescent Protein ( EYFP ) for functional assessment. Promoter activity was quantified by fluorescence under induction and repression conditions in the engineered yeast cells. Among the promoters analyzed, only the LAD promoter led to high fluorescence under induction conditions, and low under repression. Yeast cells harboring the LAD promoter showed mostly an increasing fluorescence with culture time. Moreover, dose-response analyses with arabinose from 0.05 to 30 g/L indicated that only 2.5 g/L arabinose is needed to achieve maximum fluorescence from the yeast cells. Finally, we performed a two-stage culture of X. dendrorhous harboring the LAD promoter and arabinose as inducer for prospective novel practical applications. Overall, we believe that the LAD promoter is a promising tool for fine-tuning of protein expression in X. dendrorhous . A native arabinose-inducible promoter is disclosed in the yeast. LAD promoter exhibits a sensitive response to the inducer. A two-stage culture of X. dendrorhous allows first the accumulation biomass, and then the expression of EYFP.