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Engineering CREB-activated promoters for adenosine-induced gene expression
John Robert Cox1, Andrea Fox1, Conor Lenahan1
1School of Chemical and Biomolecular Engineering, Georgia Institute of Technology, Atlanta, Georgia, USA.
Biotechnology Journal
|February 25, 2024
Summary
Researchers developed synthetic promoters activated by adenosine (ADO) to control gene expression in human cells. These novel CREB-activated promoters offer strong, tunable gene induction, overcoming challenges in cancer immunotherapy research.
Area of Science:
- Molecular Biology
- Immunology
- Synthetic Biology
Background:
- Adenosine (ADO) accumulation suppresses anti-tumor immune cell function via CREB signaling.
- Developing tools to modulate this pathway is crucial for cancer immunotherapy.
Purpose of the Study:
- To engineer synthetic CREB-activated promoters for precise control of gene expression in response to ADO.
- To overcome limitations in existing methods for studying ADO-mediated immune suppression.
Main Methods:
- Designed and optimized CREB-binding sites and core promoter regions for synthetic promoters.
- Tested promoter activity in human cells with varying ADO concentrations and signaling analogs.
- Investigated the impact of fetal bovine serum (FBS) on ADO stability and promoter function.
Main Results:
- Created synthetic promoters with up to 47-fold induction by ADO and minimal basal expression.
- Demonstrated synergistic gene expression with ADO and signaling analogs.
- Identified and mitigated ADO degradation by FBS, enabling physiologically relevant induction.
Conclusions:
- Engineered synthetic promoters provide a powerful tool for modulating ADO-CREB signaling in cancer research.
- Optimized experimental conditions, including FBS treatment, are essential for accurate ADO dose-response studies.
- Synthetic promoters can be further enhanced by incorporating additional transcription factor binding sites.
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