Engineered Feedback Employing Natural Hypoxia-Responsive Factors Enhances Synthetic Hypoxia Biosensors

Inadequate oxygen supply is a feature of multiple acute and chronic diseases, and hypoxia biosensors can be deployed in engineered cells to study or treat disease. Although mediators of hypoxia-responsiveness have been characterized, dynamics of this response are less understood, and there is no general approach for tuning biosensor performance to meet application-specific needs. To address these gaps, we investigated the use of genetic circuits to enhance biosensor performance through feedback, ultimately achieving both low background and amplified hypoxia-induced gene expression. To build insight into the mechanisms by which our circuits modulate performance, we developed an explanatory mathematical model. Our analysis suggests a previously unreported dual regulatory mechanism in the natural hypoxia response, providing new insights into regulatory dynamics in chronic hypoxia. This study exemplifies the potential of using synthetic gene circuits to perturb natural systems in a manner that uniquely enables the elucidation of novel facets of natural regulation.