High-Throughput Metabolic Engineering of Yarrowia Lipolytica Through Gene Expression Tuning

Strategic Insights -

New research by investigators at the Technical University of Denmark (DTU) has developed a high-throughput metabolic engineering approach called TUNEYALI, which enables the simultaneous testing of numerous hypotheses on gene expression in the industrial yeast Yarrowia lipolytica. The method, funded by Novocure Limited and the European Research Council (ERC), uses CRISPR-Cas9-based editing to replace or remove promoters of target genes, resulting in a library of plasmids that can be used to screen for desired phenotypic changes. This research has significant implications for applied strain development projects and fundamental functional genomics research.

Key Takeaways:

  • TUNEYALI is a high-throughput metabolic engineering approach that enables the simultaneous testing of numerous hypotheses on gene expression in Yarrowia lipolytica.
  • The method uses CRISPR-Cas9-based editing to replace or remove promoters of target genes, resulting in a library of plasmids that can be used to screen for desired phenotypic changes.
  • The research identified multiple transcription factors (TFs) whose regulatory changes increased thermotolerance, two TFs that eliminated pseudohyphal growth, and several TFs that increased betanin production.
  • The libraries can be shared and reused, accelerating applied strain development projects and fundamental functional genomics research.
  • The TUNEYALI-TF kit and TUNEYALI-TF library are available via AddGene under catalog numbers #1000000255 and #217744.
  • The research was funded by Novocure Limited and the European Research Council (ERC).
  • Irina Borodina, Wei Jiang, Shengbao Wang, Daniel Ahlheit, Tommaso Fumagalli, Zhijie Yang, Shreemaya Ramanathan, Xinglin Jiang, Tilmann Weber, and Jonathan Dahlin were among the researchers contributing to this study.
  • The research has significant implications for the field of bioengineering and metabolic engineering.

Statistics:

  • 56 transcription factors (TFs) were targeted in the TUNEYALI library.
  • Seven different levels of expression were tested for each TF.
  • The library was used to screen for changes in morphology, thermotolerance, or improved betanin production in Yarrowia lipolytica.
  • The genetic markup of the yeast clones with the desired phenotypic changes was determined by sequencing the inserted plasmids.

Sources:

  • NewsRx. Findings on Metabolic Engineering Detailed by Investigators at Technical University of Denmark (DTU) (High-Throughput Metabolic Engineering of Yarrowia Lipolytica Through Gene Expression Tuning). Biotech Week. July 16, 2025; p 581.
  • Proceedings of the National Academy of Sciences. High-Throughput Metabolic Engineering of Yarrowia Lipolytica Through Gene Expression Tuning. 2025;122(23).
  • National Academy of Sciences. www.nasonline.org/
  • AddGene. www.addgene.org/