Project 1: Genome-wide analysis of DNA replication fork stalling due to environmentally-induced DNA damage
Project 2: Ozone-induced responses in human volunteers: linking genomic profiles in airway cells with biological responses
Project 3: Free to Breathe, Free to Teach: indoor air quality in schools and respiratory health of teachers
Project 4: High throughput analysis of environmental and endogenous agents
Project: Genome-wide analysis of DNA replication fork stalling due to environmentally-induced DNA damage
Benzo[a]pyrene (B[a]P) and Ultraviolet (UV) radiation are ubiquitous environmental agents that damage DNA and cause cancer. During S-phase DNA replication forks are particularly vulnerable to the detrimental consequences of DNA damage. Failure to accurately replicate and repair DNA leads to many hallmarks of cancer cells including mutagenesis and gross chromosomal rearrangements. This project will test the hypothesis that DNA replication defects arising from environmental DNA-damaging agents are over-represented at specific loci in the human genome. We have devised an innovative protocol for isolating chromatin at sites where replication fork stalling occurs specifically in response to B[a]P- or UV-induced DNA damage. We will perform next-generation sequencing and Microarray-based Comarative Genomic Hybridization (CGH) to perform genome-wide analysis of replication events in occurring in the absence and presence of genotoxins. The CEHS Biostatistics and Bioinformatics core facility will provide expertise and assistance for alignment of sequence reads on the human genome and identification of sites of significant over-representation. We anticipate we will be able to identify novel DNA sequences and epigenetic marks associated with replication fork stalling due to environmental carcinogens.
Project: Ozone-induced responses in human volunteers: linking genomic profiles in airway cells with biological responses
Ozone (O3) continues to be of great public health importance, including in North Carolina, where O3 levels reach dangerous levels every year. O3-exposure results in pro-inflammatory responses in the lung, marked by influx of neutrophils, release of inflammatory mediators, and exacerbation of existing diseases, such as asthma. Analysis of gene expression profiles induced upon O3 exposure in the lung has never been done in humans. The goal of the studies proposed here is to identify biomarkers of exposure to O3 in airway cells collected from human volunteers. Specifically, we will use existing RNA samples and biological data available to us through ongoing studies in which human volunteers are exposed to O3 and airway cells are obtained before and after exposure. Using oligonucleotide arrays we will establish miRNA and mRNA signatures associated with O3 exposure and associate gene expression profiles with biological outcomes, such as neutrophil influx, for each individual. We anticipate uncovering miRNA and mRNA signatures that will yield novel insights into potential mechanisms of O3-induced adverse health effects and identify markers of susceptibility to O3-induced responses in humans.
Project: Free to Breathe, Free to Teach: indoor air quality in schools and respiratory health of teachers
Recent research suggests that teachers have the highest asthma prevalence of any occupational group in the United States. Asthma is also a leading cause of student absences from school. Due to the high number of occupants and scarce funds for maintenance, school buildings often suffer from poor indoor air quality (IAQ). In many schools, excessive moisture is a recurrent problem which increases exposure to asthma triggers such as mold, dust mites, roaches, and rodents. We will investigate the impact of relative humidity levels on asthma exacerbations among teachers at twelve public schools in North Carolina (NC). Though relative humidity can be cheaply measured and controlled, ours will be the first longitudinal study of humidity and asthma exacerbations in teachers. We will assess structural factors affecting indoor air quality (IAQ) in schools and monitor teachers’ respiratory health over time. Our study will provide locally relevant, scientific evidence of practical methods to reduce asthma triggers in schools. This research will be paired with training on sustainable IAQ practices to reduce the burden of environmental illnesses in schools through multi-agency collaboration.
Project: High throughput analysis of environmental and endogenous agents
The rapid assessment of cellular responses to different biological, environmental, and endogenous challenges is critical for understanding how these agents may increase genomic instability, disease formation and ultimately affect quality of life. One way to assess this response is to determine their influence on DNA replication and how this change in replication dynamics is dealt with by the cell. Recently, we have adopted the technique of replication analysis of DNA on extended single DNA fibers spread onto glass slides to expand our understanding of how DNA damaging agents influence DNA replication. This CEHS pilot project is designed to allow us to develop automated techniques to perform immunostaining and computer-assisted analysis of the slides, thereby decreasing analysis time and eliminating bias associated with analyzing the fibers manually. In addition, this methodology would accelerate the study of the effect of DNA damaging agents on isogenic cell lines with different defects in damage recognition, repair, and checkpoint pathways, thus increasing the depth of our understanding of the role of the various proteins involved in DNA replication and repair.
|Funded by NIEHS Grant # P30 ES010126|