An engineered epigenetic transgene switch in mammalian cells

Area of Science:

• Synthetic biology
• Epigenetics
• Mammalian cell engineering

Background:

• Cell identity in multicellular systems is regulated by epigenetic circuits.
• These circuits dictate cell-specific transcriptomes and phenotypes.
• Controlling gene expression is crucial for engineering cellular functions.

Purpose of the Study:

• To develop a mammalian epigenetic circuitry capable of switching between two stable transgene expression states.
• To demonstrate the functionality of this epigenetic toggle switch in engineered cells.
• To explore the potential of this system for artificial gene networks.

Main Methods:

• Combined two mutually repressing genes to create a toggle switch mechanism.
• Engineered Chinese hamster ovary (CHO-K1) cells with the toggle switch circuitry.
• Administered transiently two alternate drugs to induce state switching.
• Cultured engineered cells and assessed glycoprotein expression.
• Implanted microencapsulated cells into mice to evaluate in vivo performance.
• Utilized mathematical models to predict switch dynamics and expression stability.

Main Results:

• Demonstrated stable switching between two transgene expression states in CHO-K1 cells.
• Observed toggle switch-specific expression profiles of a human glycoprotein in vitro.
• Confirmed functional expression of the glycoprotein in microencapsulated cells after in vivo implantation.
• Validated predictive accuracy of mathematical models for switch behavior.
• Showcased the stability and controllability of the epigenetic toggle switch.

Conclusions:

• Developed a novel mammalian epigenetic toggle switch for controlling transgene expression.
• This system provides a robust tool for engineering artificial gene networks in mammalian cells.
• The toggle switch enables stable, switchable cell phenotypes with potential applications in biotechnology and regenerative medicine.