BRIGE - Prokaryotic Real Time Gene Expression Profiling For Toxicity Assessment And Water Quality Monitoring

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PI: Gu, AprilProposal Number: 0926284Goals: The health risk associated with the plethora of emerging contaminants necessitates a breakthrough in toxicity assessment technology for water quality monitoring. The goals of this project are: a) to explore thevalidity of a novel application of prokaryotic real time gene expression profiling for assessing toxic effects and understanding toxicity mechanisms, b) to develop a more sophisticated and informative, yet feasible andreliable toxicity assessment methodology for evaluation and identification of contaminants in water samples and c) to establish a creative and integrated education program to attract and educate personnel from various disciplines and identities, especially women and underrepresented groups, to become contributors in the environmental engineering field. The long-term goal is to build a unique interdisciplinary and translational research and educational program that specializes in applying genomic/molecular biotechnology for waterquality monitoring.Methods: A new prokaryotic real-time gene expression profiling method, using a comprehensive library (cellarray) of transcriptional fusions of Green Fluorescence Protein (GFP) to each of about 1900 different gene promoters in E. coli. k12, will be employed for toxicity evaluation and identification in water samples. The high-temporal-resolution and high-throughput measurements of cellular-level collective state geneexpression activities in the living cell will yield characteristic gene expression profiling in response to a specific compound or a mixture of compounds, depending on their toxic mechanisms and modes of action. A database of signature gene expression profiles (biomarkers) for various categories of emerging environmental pollutants will be established and the toxic mechanisms will be revealed. Multidimensional hierarchical clustering methods will be applied to identify and/or classify pollutants based on their similarity in gene expression profiles and, by extension, similarity in their underlying modes of action of toxicity. Geneexpression profiling results in real environmental samples will be compared with those obtained by conventional toxicity evaluation methods in order to confirm the validity of gene expression profiling fortoxicity assessment. The education plan is developed by integrating a series of teaching and educationoutreach activities with an interdisciplinary, experiential and multi-channel teaching paradigm that will incorporate the learning of existing knowledge in classrooms, discovering new information through researchand discovery, and applying and translating knowledge through practice and application. Highlights of theseactivities include expanding the pioneering BEST (Biotechnology for the Environment-Showcase and Training) program, through connections with various NU education identities, local organizations andcommunities, to increase the diversity of participants in the environmental engineering field, especially forwomen, minorities and people with disabilities; developing a new interdisciplinary course to attract andexpose students from other disciplines to environmental engineering and enhancing technology transfer andinformation dissemination through multi-level collaborations, Co-op program at NU and via ITRI (IndustrialTranslational Research Initiative).Intellectual Merits: This project integrates diverse disciplines of biotechnology, toxicology and environmental engineering and opens new ground for research in genomic-based toxicity assessment for water quality monitoring. We are the first to apply a prokaryotic cell-array consisting of a large number of bioluminescent GFP-transformed E. coli. strains to measure the real-time gene expression activities in response to contaminants. This approach will lead to timely and more informative toxicity evaluation results than conventional methods. It has higher sensitivity and specificity than the existing microarray-based genomic profiling approach because it adds a temporal dimension to the profiling data and therefore allows for more systematic and higher-resolution toxicity evaluation of pollutants. Moreover, the proposed method greatly improves the feasibility and cost-effectiveness of gene expression profiling for toxicity assessment due to its simpler, faster and reliable assay procedures, higher reusability and desirable flexibility for customization of the cell-array library.Broader Impacts: This research has significant impact on ensuring water quality for public health protectionand life quality improvement. Current available toxicity assessment methods for water quality monitoring are not sufficient, and they severely hamper our progress in water quality protection and improvement. This project presents a new interdisciplinary approach to meet the extremely urgent need for effective and reliable methods to detect and evaluate toxicity effects of emerging contaminants in water. The integrated interdisciplinary, experiential and multi-channel teaching and education paradigm will increase the diversityof participation in environmental engineering workforce and it will equip them with adequate knowledge and skills to tackle today's challenging environmental issues. The support for the PI's career development will help her develop as a productive scholar who benefits society by contributing to the technological and educational advancement of water quality improvement and public health protection.