Engineering a synthetic gene circuit for high-performance inducible expression in mammalian systems.
Giuliano De CarluccioVirginia FuscoDiego Di BernardoPublished in: Nature communications (2024)
Inducible gene expression systems can be used to control the expression of a gene of interest by means of a small-molecule. One of the most common designs involves engineering a small-molecule responsive transcription factor (TF) and its cognate promoter, which often results in a compromise between minimal uninduced background expression (leakiness) and maximal induced expression. Here, we focus on an alternative strategy using quantitative synthetic biology to mitigate leakiness while maintaining high expression, without modifying neither the TF nor the promoter. Through mathematical modelling and experimental validations, we design the CASwitch, a mammalian synthetic gene circuit based on combining two well-known network motifs: the Coherent Feed-Forward Loop (CFFL) and the Mutual Inhibition (MI). The CASwitch combines the CRISPR-Cas endoribonuclease CasRx with the state-of-the-art Tet-On3G inducible gene system to achieve high performances. To demonstrate the potentialities of the CASwitch, we apply it to three different scenarios: enhancing a whole-cell biosensor, controlling expression of a toxic gene and inducible production of Adeno-Associated Virus (AAV) vectors.
Keyphrases
- poor prognosis
- small molecule
- gene expression
- transcription factor
- copy number
- dna methylation
- crispr cas
- binding protein
- genome wide identification
- long non coding rna
- stem cells
- single cell
- blood pressure
- gold nanoparticles
- mesenchymal stem cells
- bone marrow
- cell therapy
- heat shock
- drug induced
- diabetic rats
- label free