A live zebrafish-based screening system for human nuclear receptor ligand and cofactor discovery

Area of Science:

• Molecular Biology
• Genetics
• Pharmacology

Background:

• Nuclear receptors (NRs) are crucial transcription factors regulating vital physiological processes.
• NRs are significant drug targets, yet many remain uncharacterized or are 'orphan receptors'.
• Existing methods for studying NR activity are limited in scope and application.

Purpose of the Study:

• To develop a versatile in vivo reporter system for monitoring human nuclear receptor activity in live zebrafish.
• To create a system facilitating the purification of NR-bound molecules for drug discovery.
• To establish a comprehensive resource of transgenic zebrafish lines for NR research.

Main Methods:

• Development of a GFP-based reporter system in zebrafish (Danio rerio) for NR activity monitoring.
• Engineering human NR fusion proteins with an affinity tag for molecule purification.
• Generation of transgenic zebrafish lines for a majority of human NRs.

Main Results:

• The reporter system accurately reflects NR activity in response to endogenous hormones and external drugs.
• Efficient purification of receptors and bound cofactors from responsive tissues was achieved.
• Quantifiable signals were obtained, demonstrating the system's robustness and sensitivity.

Conclusions:

• The developed zebrafish NR reporter system is a powerful tool for studying NR function in vivo.
• This system facilitates the identification of tissue- and isoform-specific ligands and cofactors.
• The established transgenic lines offer a valuable resource for NR drug discovery and research.