Mining novel gene targets for improving tolerance to furfural and acetic acid in Yarrowia lipolytica using whole-genome CRISPRi library.

[Display omitted] • Establishment of the first whole-genome CRISPRi library in Y. lipolytica. • Discovery of 14 novel gene targets for improving furfural and acetic tolerance. • Tolerance to the highest known acetic acid content (0.35%, v/v) in Y. lipolytica. • Tolerance mechanism entails enhanced cell division and decreased ROS level. • Tolerance to dual inhibitors during co-utilization of glucose and xylose. Abundant renewable resource lignocellulosic biomass possesses tremendous potential for green biomanufacturing, while its efficient utilization by Yarrowia lipolytica , an attractive biochemical production host, is restricted since the presence of inhibitors furfural and acetic acid in lignocellulosic hydrolysate. Given deficient understanding of inherent interactions between inhibitors and cellular metabolism, sufficiently mining relevant genes is necessary. Herein, 14 novel gene targets were discovered using clustered regularly interspaced short palindromic repeats interference library in Y. lipolytica , achieving tolerance to 0.35 % (v/v) acetic acid (the highest concentration reported in Y. lipolytica), 4.8 mM furfural, or a combination of 2.4 mM furfural and 0.15 % (v/v) acetic acid. The tolerance mechanism might involve improvement of cell division and decrease of reactive oxygen species level. Transcriptional repression of effective gene targets still enabled tolerance when xylose was a carbon source. This work forms a robust foundation for improving microbial tolerance to lignocellulose-derived inhibitors and revealing underlying mechanism. [ABSTRACT FROM AUTHOR]