Designing strong inducible synthetic promoters in yeasts

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Summary

The study presents a simple and reliable approach for constructing strongly inducible synthetic promoters with minimal leakiness in yeasts. The results indicate that leakiness can be largely mitigated by insulation, increasing operator repeats, and mutating causal operators.

Highlights

• The study demonstrates the construction of inducible synthetic promoters in yeasts with minimal leakiness.
• Insulation, increasing operator repeats, and mutating causal operators can mitigate leakiness.
• A generalizable design for creating potent iSynPs in various eukaryotic yeasts was established.
• The constructed iSynPs were used to produce two pharmaceutical proteins.
• The protein production platform using the iSynPs is characterized by flexibility in incubation conditions and carbon sources.
• The study demonstrates the rapid production of RBD protein from a SARS-CoV-2 omicron variant using the DAPG-inducible expression platform.
• The yeast-based platform can produce sufficient amounts of virus antigens for chicken immunization.

Key Insights

• The study highlights the importance of insulation in reducing leakiness in inducible synthetic promoters, demonstrating that the addition of a 1.6-kb insulation sequence can significantly reduce leakiness.
• The direct fusion of bacterial operator repeats to the TATA-box sequence is crucial for constructing strong iSynPs, and increasing operator repeats can monotonically increase fold induction.
• The study demonstrates the utility of the constructed iSynPs in producing pharmaceutical proteins, including nanobodies, and highlights the flexibility of the protein production platform.
• The DAPG-inducible expression platform can be used for the rapid production of difficult-to-express proteins, such as the RBD protein from a SARS-CoV-2 omicron variant.
• The study highlights the potential of the yeast-based platform for vaccine production against new pandemic viruses, demonstrating the production of sufficient amounts of virus antigens for chicken immunization.
• The constructed iSynPs can be used to produce single biologics via large-scale fed-batch fermentation, and the inducer-OFF system can minimize induction costs.
• The study demonstrates the importance of considering the economic viewpoint in the production of biologics, highlighting the potential of the inducer-OFF system to reduce costs.

Mindmap

Citation

Tominaga, M., Shima, Y., Nozaki, K., Ito, Y., Someda, M., Shoya, Y., Hashii, N., Obata, C., Matsumoto-Kitano, M., Suematsu, K., Matsukawa, T., Hosoya, K., Hashiba, N., Kondo, A., & Ishii, J. (2024). Designing strong inducible synthetic promoters in yeasts. In Nature Communications (Vol. 15, Issue 1). Springer Science and Business Media LLC. https://doi.org/10.1038/s41467-024-54865-z