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Record Count: 14
To sort columns alphabetically or numerically, click on the column
header (Title, Principal Investigator, Institution, City, ST, Award Code, or
Pubs).
DESCRIPTION (provided by applicant):
The overall goal of our research is to understand the molecular mechanisms of toxicity/carcinogenicity of environmental pollutants and the endogenous cellular defense systems to cope with pollutants. Drinking water contaminated with arsenic, a known carcinogen, is a worldwide public health issue. Epidemiology studies have linked arsenic exposure to human cancers, including skin, liver, lung, kidney, prostate, and bladder cancer. Arsenic can also cause cellular damage through generation of reactive oxygen species (ROS) that are even involved in the initiation, promotion, and progression of tumors. Although arsenic is a well defined carcinogen, it is not mutagenic and induces malignant transformation possibly by an epigenetic or cell signaling mechanism. Eukaryotic cells have evolved several defense mechanisms to cope with stress from the environment, one of which is the antioxidant response utilized by mammalian cells to neutralize ROS and to maintain cellular redox homeostasis. This antioxidant system is mediated through the antioxidant response element (ARE) sequence present in the promoters of several antioxidant and Phase II detoxification genes including glutathione S-transferase, NAD(P)H quinone oxidoreductase, glutamylcysteine synthetase, and heme-oxygenase. The antioxidant response system is mainly controlled by the transcription factor Nrf2. Activated by compounds possessing anti-cancer properties, the ARE-Nrf2-Keap1 signaling pathway has been clearly demonstrated to have profound effects on tumorigenesis. More significantly, Nrf2 knockout mice display increased sensitivity to chemical toxicants and carcinogens and are refractory to the protective actions of chemopreventive compounds. Therefore, we hypothesize that activation of the ARE-Nrf2-Keap1 pathway acts as an endogenous protective system against arsenic-induced toxicity and carcinogenicity. The following specific aims are intended to further elucidate the mechanism of Nrf2-activation in protection from arsenic-induced toxicity/tumorigenicity. This knowledge can potentially serve the scientific and medical community in our objective to create novel chemopreventive agents with increased specificity and efficacy, which will have broad impact on human health worldwide. We propose to (1) determine the protective role of the ARE-Nrf2- Keap1 pathway in arsenic-induced toxicity and carcinogenicity, (2) define the molecular mechanisms of activation of the ARE-Nrf2-Keap1 pathway by arsenic, and (3) define the protective role of the ARE-Nrf2-Keap1 pathway in arsenic-induced toxicity and tumorigenicity using the Nrf2 knockout mouse model.
DESCRIPTION (provided by applicant): Hexavalent chromium (Cr(VI)) is a known human lung carcinogen. Millions of workers are exposed to Cr(VI) worldwide. The most recent paradigm proposes that the chromate compounds that are moderately water-soluble are more carcinogenic than the chromates that are either insoluble or water-soluble; however, it is not understood why this is so. Most of the data on how chromium damages DNA and causes DNA mutations has come from either the insoluble chromates or the water soluble chromates, not from the moderately soluble chromates that may be the most carcinogenic. The moderately soluble chromates are zinc chromate, strontium chromate and calcium chromate. Several crucial pieces of information are lacking regarding the activity of these moderately soluble chromates. No mutagenicity studies have been carried out with these compounds in human lung epithelial cells, which are the target cells for tumor formation. The types of mutations that these compounds cause have not been characterized in any cell line. The relative mutagenic potency of these chromates is not known. The extent to which these compounds dissolve outside of cells or enter cells as particulates is not known. The involvement of the zinc, strontium and calcium counterions in chromate toxicity is not known. The goals of the current proposal are to explain (1) how the moderately soluble Cr(VI) compounds enter cells and (2) if the mutations caused by the moderately soluble chromates differ from those caused by the soluble and insoluble chromates. The first aim of this proposal will apply the techniques of inductively coupled plasma mass spectrometry, laser scanning confocal microscopy, transmission electron microscopy, and scanning electron microscopy to determine how the moderately soluble chromates enter cells. The second aim of this proposal will measure and characterize mutations at the hypoxanthine (guanine) phosphoribosyl transferase (hprt) locus caused by the moderately soluble chromates in human lung epithelial cells and will compare mutation frequency and identity with mutations caused by the soluble and insoluble chromates. Data from these experiments will determine the relative mutagenic potency of these chromates, and will give insight into mechanisms of action, i.e., involvement of counterion and possible DNA lesions responsible for the mutations. Data from this proposal will provide mode of action information that will be necessary for thorough human risk assessment. The purpose of this work is to determine how chromium(VI) causes cancer. The goals of this proposal are to explain (1) how the moderately soluble chromium(VI) compounds enter cells and (2) if the mutations caused by the moderately soluble chromates differ from those caused by the less carcinogenic soluble and insoluble chromates. Understanding the mode of action of these chromates will provide a foundation for human risk assessment.
The vast majority of Superfund sites in the U.S and the Baltimore/Chesapeake Bay area contain mixtures of
organic and inorganic compounds that contaminate underlying aquifers. The environmental fate of these
contaminants, and ultimately human exposure to them, is governed primarily by their interactions with
microorganisms, which?individually or as a community?drive the process of in situ bioremediation.
Whereas single pollutant/microorganism interactions can be determined easily in the lab, no satisfactory
tools exist for predicting the fate of mixtures in the environment. Our long-term goal is to improve the
success rate of bioremediation at sites containing complex chemical mixtures by using in situ microcosm
array (ISMA) technology. The ISMA is a field-deployable, miniaturized laboratory consisting of a large
number of small microcosms arranged in parallel. Upon deployment, incubation, and retrieval from a
groundwater well, the ISMA can be analyzed to reveal the impact of mixture components on the rates of
Dollutant degradation and on the structure and function of microbial communities. We hypothesize that the
ISMA can aid in the design of bioremediation strategies because: (1) individual ISMA microcosms can be
amended with multiple test substances to elucidate the effect of mixture components on microorganisms; (2)
the response of microorganisms to presented compounds manifests itself as changes in biomass,
community structure and function; and (3) these changes can be detected conveniently by biochemical,
genetic and proteomic strategies. Based on the above observations, the specific aims of the project are to:
1. Determine the reproducibility and discriminatory power of the ISMA technology. We will explore how
varying concentrations of inducers and co-contaminants in synthetic groundwater modulate the expression of
the dioxin dioxygenase (DDase) of Sphingomonas wittichii Strain RW1, by using a large number of replicates
in conjunction with semi-automated, high-throughput proteomic mass spectrometry.
2. Demonstrate in controlled laboratory conditions how the ISMA can reveal additive, synergistic, and
antagonistic effects of mixture components on microbial communities. We will use defined mixtures of
chemicals (e.g., dioxins, PCBs, PAHs, tolualdeyhyde, Cd, Cr, Co, Pb, Hg, Zn, Ni) and bacteria (five
bioremediation agents) to study these effects in various permutations.
3. Evaluate the utility of the ISMA technology in simulated field conditions. We will conduct ISMA laboratory
experiments using nonsterile groundwater, sediment, and natural microbial communities.
4. Deploy the ISMA device at a Maryland Superfund site. We will assess the utility of the new technology in a
field demonstration study by examining survival of and DDase expression by Strain RW1 in situ, and by
elucidating the impact of this introduced bacterium on the indigenous microbial community at the site.
DESCRIPTION (provided by applicant):
Metalliferous mine tailings in arid regions pose a significant health risk to proximal populations because they are prone to wind-borne dispersion and water erosion. The problems are extensive and persistent as impacted sites lack normal soil stabilization. Phytostabilization is the revegetation of mine tailings to ameliorate these issues with the goal of root zone metal accumulation to avoid metals from entering the food chain through above-ground biomass. The role of plant roots and microbes in promoting mineral dissolution-precipitation reactions and associated metal sequestration is an active area of research, but little is known about reaction trajectories and changes in particle-scale metal speciation of plant-tailings systems, owing largely to their geochemical heterogeneity and microbial complexity. Since the form or speciation of a metal controls its bioavailability and toxicity, research that probes coupling between metal speciation and microbial dynamics in response to phytostabilization is needed.
The overarching goal of the proposed work is to identify multi-scale process-links between biological structure and contaminant geochemistry during phytostabilization of mine tailings. The four Specific Aims are: (i) to deduce the dependence of metal(loid) molecular environment on particle-specific weathering processes; (ii) to assess the spatial correlations between biogenic (and geogenic) weathering products and specific microbial cells and biofilms; (iii) to relate these direct observations of solid phase biogeochemistry with time- and depth-resolved measurements of tailings pore waters (focusing on the mobility and bioavailability of metal contaminants); and (iv) to measure the influence of solid and solution phase dynamics [(i) through (iii)] on the evolution of tailings microbiology and geochemistry both in the rhizosphere and in the bulk tailings over the course of phytostabilization. Embedded within these objectives is the additional goal of statistically integrating the nano- to macro-scale "geo" and "bio" information gained to better understand the phytostabilization process and possible outcomes in terms of exposure and toxicity risks associated with tailings sites in arid environments. Biostabilization will be probed over a 27 month mesocosm experiment using an array of advanced tools that can interrogate the complex associations of roots, microbes, minerals and metals at high spatial resolution. A time series of bulk and micro-focused X-ray spectroscopic, molecular biology/microbial ecology, and aqueous geochemical data will be generated for analysis of coupled processes that control the local contaminant environment. To assess how these process-links affect the larger goal of metal stabilization, we will coordinate our "bio" and "geo" observations so that they probe identical locations, together traversing molecular to macroscopic (mesocosm-level) scales. This research is both timely and necessary as growth in the US Southwest is exploding and communities are being developed in closer proximity to such tailings sites.
DESCRIPTION (provided by applicant)
The theme that unites the research activities of the Southwest Environmental Health Sciences Center (SWEHSC) is the need to understand the mechanisms by which exposures to environmental agents contribute to human disease. Identifying factors that contribute to human diseases that arise as a consequence of environmental exposures is a prerequisite for the development of strategies designed to minimize both the environmental exposure, and the adverse effects of such exposures. The SWEHSC is a dynamic organization that promotes collaborative, interdisciplinary research, with an emphasis on the detection and prevention of environmentally-mediated diseases. Members of the SWEHSC are organized into three Research Cores (RC) with overlapping spheres of interest; Mechanisms of Environmental Chemical Toxicity (RC1), Pulmonary Toxicology and Lung Disease (RC2), and Chemical-Chromatin Interactions (RC3). The Research Cores provide the scientific foundation that provides the basis for strategies designed to treat or prevent environmental diseases. The research efforts of SWEHSC are complemented by five Facility Cores (FC); Cellular Imaging, Genomics, Proteomics, Synthetic Chemistry, and Bioinformatics. The Facility Cores offer state-of-the-art instrumentation and expertise to assist investigators in developing and utilizing cutting-edge technologies. The Administrative Core provides enrichment programs, including seminars, workshops, an annual Science Fair, newsletters, and supports the activities of the Internal and External Advisory Boards. The SWEHSC is also actively involved in promoting innovative research through the Pilot Project Program, and in the recruitment of new investigators. The Community Outreach and Education Program (COEP) offers environmental health science and toxicology based educational programs that are geared to students (K-12), health professionals, and the public. Through the COEP, the SWEHSC provides service to health organizations, government agencies, and the private sector. Three major inter-dependent goals will provide the focus for the next 5 years of support. The SWEHSC will (1) Facilitate the utilization and integration of" global" systems-centered toxicological approaches (proteomics, toxicogenomics, SNP's, cell and tissue imaging) to basic environmental health science research questions. (2) Facilitate the extension of basic research discoveries into the clinical and public health arenas. SWEHSC faculty, in collaboration with faculty in the Colleges of Public Health, Medicine, Pharmacy, and Nursing, will focus in the general area of novel biomarkers (SNP's, proteins, protein adducts, etc) of susceptibility, particularly within minority populations (Hispanic, Native American). Emerging border health initiatives will play an integral role in developing these new areas of emphasis. (3) Initiate and expand collaborations with sister Centers of Excellence on the University of Arizona Health Sciences Center campus (e.g., Arizona Respiratory Center, The University of Arizona Sarver Heart Center, Arizona Cancer Center, Arizona Center on Aging, Steele Memorial Children's Research Center and the BIOS Institute). This last goal is synergistic with the objective of extending basic research discoveries into the clinical and public health arenas.
DESCRIPTION (provided by applicant)
The title of this proposal captures its essence, 'Hazardous Waste Risk and Remediation in the Southwest'. The University of Arizona SBRP renewal application builds on the achievements of the previous grant. The theme of this Program is to support development of a risk assessment process for metal and organic contaminants through toxicologic and hydrogeologic studies and through development of innovative remediation technologies. The application emphasizes hazardous waste issues in the Southwestern U.S. (and Mexican Border) due to the unique arid nature of this environment. However, the results of the studies are not limited to the Southwest since the main toxicants being examined, As and halogenated hydrocarbons, are ubiquitous throughout developed countries. The Program consists of 10 research projects-five biomedical projects and five environmental sciences projects. Many of the projects are collaborative involving multiple disciplines. The biomedical projects are examining the mechanism of As toxicity in target tissues, factors that affect the susceptibility of populations to As-induced toxicity, and the effects of As and TCE on organ development. The environmental sciences projects are investigating how hazardous wastes (As, TCE/perchloroethylene [PCE]) can be optimally characterized for remediation and innovative techniques for containing or degrading these contaminants in an arid Southwest environment. These research projects are supported by 5 cores that administer the Program, translate the results to the stakeholders, provide research services, promote unique outreach efforts to Mexico, and support graduate student training. This project will contribute to our understanding of toxicology and remediation of hazardous wastes nationally and internationally.
DESCRIPTION (provided by applicant): The Navajo Nation is home to the largest Native American tribe in the U.S. In the mid 1940's active uranium (U) mining began in the Four Corners region of the southwest U.S. where the Navajo Nation is located. U mining continued for 4 decades until the U ore market collapsed. There are over 1000 unremediated U mines on the Navajo Nation and having not closed these properly has led to widespread U contamination of the soil and water. According to the U.S. EPA there are scores of water sources used by Navajo people for drinking and household use that have U levels that exceed the safe drinking water limit of 30 ¿g/L. There are many health problems known to arise from ingesting U containing water but to date all have to do with the toxicity of U as a heavy metal. Recently we discovered that U, much like other heavy metals, has estrogenic activity. We have found that U, at equimolar concentration to diethylstilbestrol (DES), elicits estrogenic responses in the reproductive tract of female mice and in tissue culture stimulates human breast cancer cell proliferation. In recent experiments we have found that in utero exposure to DES or U causes developing pup ovaries to overproduce androgen compared to ovaries from pups whose dams drank tap water. Based on this new data we propose the hypothesis that in utero U exposure permanently alters the programming of the developing ovary so that it produces more androgen that over a lifetime contributes to premature ovarian failure and metabolic dysregulation. This hypothesis will be tested in two aims. First, we will determine if the hyperandrogenism observed in ovaries from 35-38 day old mice exposed in utero to DES or U is permanent by examining ovaries and analyzing production of androgen in tissue culture from mice up to 18 months of age. And, we will determine if androgen blood levels are increased in these mice during the LH surge of the estrus cycle. In the second aim we will determine if persistent hyperandrogenism contributes to accelerated ovarian failure by short circuiting folliculogenesis. Finally, we will analyze the global effects of lifetime hyperandrogenism by analyzing onset of insulin resistance, hyperglycemia, hypertriglyceridemia and deposition of truncal fat. Our overall goal is to test the hypothesis that in utero exposure to an endocrine disrupting estrogenic chemical, U, leads to increased risk of developing type 2 diabetes and ultimately heart disease. The prevalence of these two chronic illnesses has sky rocketed in the Navajo people in the last several decades. Growing scientific understanding has revealed that the uterine environment as a function of the mother's exposure to environmental chemicals can lead to permanent differences in the developing child that leads to as an adult greater risk for developing diabetes and heart disease.
DESCRIPTION (provided by applicant): Women are exposed daily in the workplace, as well as the environment (cigarette smoke, automobile exhaust) to chemicals that can damage small pre-antral (primordial) ovarian follicles. Damage to these follicles can result in early ovarian failure (menopause). Because menopause is associated with a variety of health disorders, this represents a plausible health risk. Dosing of mice and rats with the occupational chemical, 4-vinyl-cyclohexene diepoxide (VCD) destroys primordial follicles and early ovarian failure can be caused in rodents by repeated dosing. On-going mechanistic studies in rats have helped characterize VCD as a model chemical for the study of xenobiotic-induced destruction of primordial follicles. However, the exact mechanism, which initiates oocyte degeneration, remains unknown. Thus, it is proposed here to expand the base of mechanistic information obtained with VCD to further identify those mechanisms. Because a variety of environmental chemicals are known to destroy primordial follicles, VCD-related information will prove applicable to chemicals that are sources of greater exposure in the environment. Therefore, a comparison between selected parameters of VCD-induced ovotoxicity and another environmental chemical, 9,10-dimethylbeznanthracene, DMBA, present in cigarette smoke and automobile exhaust will also be made. The hypothesis to be tested is that VCD causes destruction of primordial follicles by upregulation of pre- and post-transcriptional intracellular pathways and DMBA initiates similar events. The Specific Aims are to: 1) identify and characterize gene expression directly regulated by VCD dosing, 2) dissect signaling pathways involved in VCD-induced ovotoxicity 3) compare selected signaling events initiated by DMBA with those of VCD, and 4) characterize the ability of ovarian compartments to bioactivate and detoxify VCH, VCD, and DMBA. Specifically investigating the mechanism(s) by which VCD damages ovarian primordial follicles and comparing this with DMBA will provide a greater ability to predict potential risks for early menopause from environmental exposures in women. This greater awareness will lead to an appreciation of the global impact of the environment on age of menopause in women
DESCRIPTION (provided by applicant):
The ability to quickly and reliably detect chemical toxicants in air is critically important for health risk assessment, for better understanding the role of gene-environment interactions in human diseases, and for health disparities research. Current detection of chemical toxicants relies on bulky and expensive spectroscopic and chromatographic techniques that require considerable maintenance and operator expertise, which are not practical for continuously monitoring various chemicals at multiple locations. Many portable devices have been proposed and developed, but they have different limitations ranging from low selectivity, insufficient sensitivity, limited scope and high costs. The present project brings together a joint effort involving chemical sensor researchers at Arizona State University (ASU), toxicologist at University of Arizona (UA), R&D scientists and engineers at Motorola and field testing experts at Arizona Division of Occupational Safety & Health (ADOSH) to build, validate and test a powerful wearable sensing system. The sensor technology is built upon a novel microfabricated tuning fork array sensor platform invented at ASU and wireless sensor technology developed at Motorola. The project will leverage on the expertise and resources gathered for an on-going collaborative R&D effort on wireless chemical sensors between the ASU and Motorola team. The goal is not only a wearable sensor system for quick, accurate and reliable detection of chemical toxicants, but also an affordable, easy-to-upgrade and user friendly product for population studies.
DESCRIPTION (provided by applicant): The goal of this Phase II Small Business Innovative Research (SBIR) application is to create a series of educational materials for the National Institute of Environmental Health Sciences (NIEHS) entitled Powers of Inquiry: Using Image Analysis to Explore Environmental Health Science. The ten lessons to be published by the project will employ four advanced technology tools-NASA Image2000 (NI2K), ArcExplorer Java Edition for Education (AJEE), Imaged, and Weblmage-to lead middle and high school students through inquiry- based explorations often case studies related to the mission of NIEHS. In Powers of Inquiry, students will use NI2K, AJEE, ImageJ, and Weblmage to display and enhance digital images from research being conducted by NIEHS-supported and other scientists. With these tools and the support provided by the Powers of Inquiry materials, students will strengthen their science and technology skills, become better consumers of scientific data, and develop an appreciation for the role that NIEHS plays in protecting the health of the nation. A "powers of ten" format will organize the lessons in Powers of Inquiry. Each lesson in the series will zoom in on environmental health issues, starting at the planetary level and moving through lessons about the stratosphere, coastal regions, communities, neighborhoods, homes, human organs, microscopic organisms, and human tissue cells to arrive at a lesson on the benefits and risks of molecule- sized nanotechnologies. The educational goal of the Powers of Inquiry materials will be to introduce environmental health science to middle and high school students with an engaging visual medium that involves them in inquiry-based activities supporting accomplishment of state and national standards for science, mathematics, technology, and reading education. The materials will be developed by Science Approach, a for-profit organization founded by staff from the Center for Image Processing in Education (CIPE). CIPE, a well-respected nonprofit provider of instructional materials and professional development services to teachers, has a successful track record of developing and commercializing instructional materials similar to Powers of Inquiry.
Crisp Terms/Key Words: educational resource design /development, clinical research, interactive multimedia, digital imaging, human subject, health science research, field study, environmental health, secondary school, elementary school, computer program /software
DESCRIPTION (provide by applicant)
The Toxicology Training Program at the University of Arizona has a long-standing reputation for producing many successful Ph.D.s. Graduates are now key players in academia, industry, and government. In response to current and future demand for qualified graduates in the environmental health sciences, the investigators have enhanced their systems-based Toxicology training with an emphasis on cellular and molecular mechanisms that incorporate genomics and proteomics approaches. The cutting-edge interdisciplinary research programs of 21 training grant faculty members, state-of-the-art technologies developed through the Southwest Environmental Health Sciences Center (SWEHSC) and translational approaches undertaken by the NIEHS Superfund Program provide an exceptionally stimulating environment for the training of graduate students and postdoctoral fellows. The Research and Facility Cores supported by the SWEHSC extend the training environment from a single laboratory-oriented domain into a multidisciplinary experience strongly supportive of interactive and collaborative research. The university provides financial support for first year Ph.D. students in the Graduate Programs of Pharmacology and Toxicology, Physiological Sciences, and Cancer Biology, resulting in a large pool of qualified candidates for competitive selection of pre-doctoral trainees. Pre-doctoral training is achieved through a combination of coursework, laboratory research, and supplemental enrichment activities. Postdoctoral trainees have ample opportunities to participate in innovative research programs and to develop their professional skills in oral and written communication and in supervision. Over the past five years, the investigators have generated six new graduate courses: Molecular Toxicology, Toxicogenomics and Proteomics, Advanced Toxicology, Environmental Toxicology Colloquium, Ethics, and Scientific Writing. The curricular changes parallel the evolving expertise of the Training Grant Faculty in genomics and proteomics. They have recruited five senior (Professor) and three junior (Assistant Professor) faculty into the Training Grant, which significantly enhances strength in an evolving theme of molecular toxicology training. The request for continuation of NIEHS support is validated by the highly successful nature of their program, the clear demand for their graduates, the increasing number of students interested in toxicology, institutional commitment, strong and well-funded research programs of the faculty, and the excellence of the training environment.
DESCRIPTION (provided by applicant)
The University of Arizona (U of A) proposes to establishment of a Human Genes and the Environment Research (HuGER) Training Program. Development of the HuGER training program has been guided by several important realities. Most relevant to the HuGER TG is recognition of the fact that in the coming decades, a more precise determination of the influence of environmental exposures within a given genetic background on disease processes will be required to significantly improve our ability to predict, detect, treat and monitor disease progression and disease response. In addition it is increasingly clear that epigenetic status will emerge as a critical process that is modulated by environmental exposures, leading to the adverse or beneficial manifestation of that exposure. The HuGER will build upon three inter-disciplinary pre-, and post-doctoral training programs integral to the creation of a successful multi-disciplinary training program that trains scientists in environmental genomics/genetics. An (i) NIEHS supported inter-disciplinary training program in Toxicogenomics and Toxicology, an (ii) NSF Interdisciplinary Graduate Education and Research Training (IGERT) Program in Evolutionary, Functional, and Computational Genomics, and (iii) a Graduate Interdisciplinary Program (GIDP) in Statistics provide the foundation for the evolution of this multi-disciplinary initiative. The HuGER curriculum has been created specifically to address the unique requirements of a multi-disciplinary training program, the cornerstone of which includes two new courses, redesigning additional courses, "industrial" research rotations, and an emphasis on the development of competent and effective communicators. This is especially important for the new generation of scientists who will need to communicate effectively across multi-disciplinary boundaries. The training environment at the U of A also provides trainees with access to appropriate contemporary computing and state-of-the-art technologies. The Training Program Faculty consist of a core of 17 scientists, from 10 departments, with active research programs in the areas of (i) the environmental and public health sciences and engineering, (ii) population and functional genomics/genetics, and (iii) computational biology and statistics/bioinformatics. Six principal units are participating in the HuGER Training Program: [1] the College of Agriculture and Life Sciences; [2] the College of Engineering; [3] the College of Medicine [4] the College of Pharmacy; [5] the College of Public Health; and [6] the College of Sciences. Five of these Colleges participate in the BIO5 Institute which brings together scientists from disparate disciplines to solve complex biological problems. The Associate Director of this HuGER application, Dr. Vicki Chandler, is the Director of BIO5. To ensure the program is known for its multidisciplinary emphasis, it will be administratively housed within BIO5.
DESCRIPTION (provided by applicant): Uranium has been mined for many years and used for fuel for nuclear reactors and materials for atomic weapons, ammunition, and armor. The radioactivity of uranium has been linked to the development of lung, kidney cancers, and leukemia. Little is known about the direct chemical genotoxicity of uranium. The purpose of the research is to gather more information about the chemical genotoxicity of uranium. The overall aims are to (1) identify the possible mechanisms induced by the chemical toxicity of uranium, (2) investigate the chemical effects of altered gene expression in HBE-16 cells, induced by exposure to depleted uranium as uranyl acetate, and (3) to determine the role of the altered genes in uranium carcinogenesis. The use of cDNA microarray to profile the gene expression of uranium-transformed cells in vitro will provide a picture as to what genes are being altered and pose questions regarding possible mechanisms of carcinogenesis. It is hypothesized that uranium will alter genes responsible for apoptosis and oncogene induction. Understanding how uranium reacts with DNA is important to better understand how this carcinogen induces cancer and to help to elucidate mechanisms of lung and other cancers in people exposed to uranium.
DESCRIPTION (provided by applicant): We propose to develop a novel general methodology for studying intra-molecular conformational changes occurring in biological macromolecules in a single cell. Our novel approach is based on the combination of Fluorescence Correlation Spectroscopy (FCS) with Fluorescence Resonance Energy Transfer (FRET), and will allow the study of a small number of molecules at the time in individual living cells. We will use a simple DNA hairpin as a "proof of principle" system to validate our novel methodology. The completion of this research will open up new avenues of research for the investigation of other dynamic biochemical processes in the cell. Although current technologies allow the observation of a single fluorescent molecule at the time, the extremely low signals obtained in such measurements limit their application to the study of slow process in solution. Fluorescence Correlation Spectroscopy (FCS), has recently emerged as an alternative to study the dynamics of processes in a wide range of timescales (¿s-s). In this technique, which has been successfully applied in solution, the fluctuations in fluorescence intensity are analyzed statistically to obtain dynamic information of the system. However, the applicability of FCS to reactions occurring in biological samples is limited by the fact that the fluctuations caused by the reaction one wishes to study are coupled to the fluctuations caused by diffusion of the macromolecule in and out of the confocal volume. Thus, the current methodologies require that the diffusion properties are evaluated in an independent experiment using a reference sample. To overcome this issue, we propose a new methodology that involves the simultaneous measurement and analysis of the fluctuations in intensity of a donor- acceptor FRET pair. We performed a preliminary theoretical study that shows that the auto (donor-donor and acceptor-acceptor) and cross-correlation (donor-acceptor) functions can be analyzed in a way that uncouples diffusion from kinetics. In this way, the dynamics of biochemical reactions will be able to be studied in environments where diffusion is difficult or impossible to characterize. We will study the opening-closing kinetic rates of a DNA hairpin using our novel FCS-FRET method. We propose to start by studying this system in solution to validate the method, and then optimize the methodology in more complex environments such as vesicles and cells. The ability to study a single cell is critical in order to understand processes like aging, and diseases like cancer. These processes are originated in a single cell due to often unknown environmental and chemical stresses. Thus, in order to understand how a given type of stress triggers the formation of an anomalous cell, it is necessary to follow the fate of the progeny of individual cells as a function of time. Furthermore, the understanding of conformational dynamics in living cells is critical to understand all its biological functions. For instance, gene accessibility to the cell's protein machinery depends on the conformational dynamics of DNA-protein interactions in chromatin. Understanding chromatin dynamics is a key step in the process of elucidating the mechanisms of development, differentiation and cancer.
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