A live-cell biosensor for real-time monitoring of cortisol dynamics

About the Project

This project will develop the first genetically-encoded biosensor for real-time monitoring of cortisol dynamics in living cells. By combining protein engineering, molecular biology, and confocal imaging, we will enable spatiotemporal analysis of cortisol signalling with unprecedented precision. The tool will transform endocrine research, revealing mechanisms of stress and metabolic regulation, and guiding therapeutic strategies for cortisol-related disorders.

Background.

Cortisol is a central glucocorticoid hormone regulating metabolism, immune responses, cardiovascular function, and the body’s response to stress. Dysregulation of cortisol signalling is implicated in a wide range of disorders, including Cushing’s syndrome, Addison’s disease and depression. Despite its crucial role, our understanding of how cortisol acts dynamically within cells and tissues is extremely limited. Current approaches largely rely on static blood measurements or endpoint assays, which fail to capture rapid, tissue-specific fluctuations or subcellular hormone distribution. Developing a tool to monitor cortisol dynamics in real time and at cellular resolution would transform our ability to study hormone regulation and provide new insights into disease mechanisms. Genetically-encoded fluorescent biosensors have revolutionised the study of ions and small molecules, enabling researchers to visualise signalling events in live cells with high spatiotemporal precision. However, no such tool currently exists for cortisol, leaving a major gap in endocrine research.

Aims and objectives.

This project aims to develop a genetically-encoded biosensor, enabling real-time monitoring of cortisol activity in living cells. The proposed work will combine molecular biology, protein engineering and live-cell imaging to provide a molecular tool for studying glucocorticoid signalling with unprecedented resolution.

The specific objectives are:

1. Design and engineer a cortisol biosensor by fusing the ligand-binding domain of the glucocorticoid receptor (GR) to fluorescent reporters optimised for real-time detection.
2. Perform in vitro characterisation of biosensor performance, including ligand sensitivity, specificity against related steroids, dynamic range, and response kinetics.
3. Validate biosensor function in live cells using confocal imaging.

Research plan.

This interdisciplinary project combines protein engineering, biophysics and confocal microscopy to investigate cortisol signalling dynamics and their role in health and disease.

• Biosensor design. The biosensor will be engineered by fusing the glucocorticoid receptor ligand-binding domain to a fluorescent reporter. We will optimise sensitivity, dynamic range, binding affinity and selectivity, ensuring a robust response to physiologically relevant cortisol concentrations.
• In vitro characterisation. Recombinant proteins will be expressed and purified for measurement of cortisol binding affinity, specificity and response kinetics.
• Live-cell functional assays. The biosensor will be expressed in cultured mammalian cells, enabling the measurement of real-time cortisol dynamics in situ. Confocal microscopy will be used to monitor subcellular localisation and distribution. Validated biosensors will be applied to study physiological and pathological cortisol signalling.

Outcome.

This project will deliver the first genetically encoded biosensor capable of reporting cortisol signalling in real time. It will provide quantitative insights into the spatial and temporal dynamics of hormone regulation. The biosensor is expected to drive advances in endocrine biology by enabling unprecedented precision in studying cortisol function.

Student training and development.

The successful candidate will receive comprehensive, multidisciplinary training in areas spanning protein engineering and live-cell imaging:

• Molecular cloning and protein engineering – design, optimisation, and in vitro characterisation of genetically-encoded biosensors.
• Fluorescence-based imaging –confocal microscopy in live cells.
• Cell biology and live-cell assays – investigation of real-time signalling dynamics in mammalian systems, linking molecular function to cellular outcomes.
• Data analysis and scientific communication – interpretation of complex datasets, preparation of manuscripts and conference presentations.