Toxin-antitoxin based transgene expression in mammalian cells

Area of Science:

• Biotechnology
• Molecular Biology
• Cell Biology

Background:

• Recombinant gene expression in mammalian cells is often limited by transgene integration site effects and epigenetic factors.
• Achieving stable, high-level transgene expression is crucial for various biotechnological applications.

Purpose of the Study:

• To develop a method for achieving and stabilizing high transgene expression in proliferating mammalian cell cultures.
• To establish a drug-free system for enhancing and maintaining recombinant protein production.

Main Methods:

• Utilized Chinese Hamster Ovary (CHO) cell lines conditionally expressing the toxin kid.
• Co-expressed the antitoxin kis with transgenes of interest, leveraging the growth advantage conferred by antitoxin expression.
• Cultivated pools of transfectants in the presence of the toxin to enrich for high-expressing cells.

Main Results:

• Transgene expression increased up to 120-fold over several weeks in cultures expressing the toxin.
• Cells with strong antitoxin expression were enriched due to a growth advantage.
• Transgene expression decreased when toxin expression was absent.

Conclusions:

• The co-expression of a transgene with its corresponding antitoxin in toxin-expressing cells provides a robust method for high and stable gene expression.
• This approach enables drug-free selection and enrichment of cells with superior transgene productivity.
• The described method offers a simple and effective strategy for generating overexpressing cell lines for recombinant protein production.