(293g) Novel Bacterial Biosensors for the Discovery of Subtype-Selective Thyroid Receptor Modulating Drugs

Thyroid hormone receptors (TRs) belong to the nuclear hormone receptor superfamily, and play a critical role in heart metabolism and adipogenesis through thyroid hormone signaling. Two subtypes of TR, α and β, exhibit obvious organ preference and are thought to play different roles in thyroid-based metabolic regulation. The discovery of subtype-selective therapeutics for the TRs is therefore a critical goal for this important drug target. This, as well as high-throughput detection of TR-targeting endocrine disruptors, calls for a subtype-distinguishable biosensor platform. Our lab has developed a modular generic biosensor where a desired nuclear hormone receptor (NHR) gene is fused to an engineered intein and thymidylate synthase (TS) reporter enzyme. Expression of this fusion in TS deficient E. coli leads to an NHR ligand-dependent growth phenotype. In the present work, two new bacterial biosensors for thyroid hormones and analogues were constructed by insertion of either the TR α 1 or TR β1 genes. These new biosensors show phenotype dependence to the natural TR ligand, 3,3',5'-triiodo-L-thyronine (T3). Further, a variety of synthetic ligands were found to cause a differential responses in the TR α1 and TR β1 sensors, suggesting these sensors are capable of distinguishing subtype-selective ligands. In addition, the sensors were able to distinguish agonist from antagonist compounds via the promotion or inhibition of cell growth. Finally, the half-maximal effect concentrations (EC50) for some ligands were measured and found to be consistent with their reported affinities to the TRs. Thus our results show that these TR α1 and TR β1 biosensors can act as a powerful tool for the discovery of subtype-selective drugs for TR-related disorders.