Development of Animal-Based Bacterial Biosensors for the Detection of Estrogenic Compounds
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Date
2015-05
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The Ohio State University
Abstract
Estrogenic compounds are an important class of chemicals that have the ability to modulate the function of the nuclear hormone receptor, estrogen receptor beta (ERβ). These compounds can be found naturally in the environment or chemically synthesized, and have been linked to health risks ranging from altered sexual development to breast cancer. Previous work in this lab resulted in an Escherichia coli cellular biosensor comprised of an engineered protein scaffold with an inserted human ERβ ligand-binding domain fused to a thymidylate synthase reporter enzyme. Through growth in thymineless media, the human ERβ biosensor can distinguish between estrogen agonists and antagonists based on the resulting E. coli growth phenotype. Preliminary results using several animal-based ERβ bacterial biosensors (cow, zebrafish, and rat) revealed species-specific responses to ligands and identified chlorodecone as an ER antagonist and dioctyl phthalate and Bisphenol B as weak ER agonists. This research sought to extend the ERβ biosensor method to include sentinel animal species, such as the zebra finch and wood frog, which serve as early indicators of the presence of environmental toxins. Using bioinformatics and molecular cloning techniques, novel sentinel species-based ERβ biosensors were created and validated against known strong and weak estrogens using the high throughput bacterial assay method. Higher sensitivity analogs of four animal biosensors were also constructed and exhibited greater than 10-fold increases in ligand response sensitivity. Unique responses to weak and strong estrogens were also observed for biosensors across a range of vertebrates (mammals, fish, birds, and amphibians), suggesting species-specific susceptibility for certain endocrine disruptors with greater deviations observed for weaker estrogens. In addition, biosensors selective for TRβ and ERα were constructed, exhibiting the ability of the ERβ biosensor to be easily modified for other pharmaceutically relevant nuclear hormone receptors. Adaptation of the ERβ biosensor to sentinel species, in particular, will enable the prevention and treatment of diseases through early detection of estrogenic compounds in the environment and expand the bacterial biosensor library. These animal ERβ bacterial biosensors will also provide a broader understanding of how drug response deviates across species in pre-clinical development.
Description
Denman Undergraduate Research Forum (2nd place in the Engineering Division) 2013
Outstanding Undergraduate Award for Research Excellence 2014
Outstanding Undergraduate Award for Research Excellence 2014
Keywords
Estrogen, Bacterial Biosensor, Sentinel species, Endocrine disruptors