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Principal Investigator: Kenneth S. Ramos
Organization: TEXAS A&M UNIVERSITY HEALTH SCIENCE CTR
Fiscal Year: 2024
Award: $643,873
Funding agency: National Institute of Environmental Health Sciences
Abstract
Functional genomics studies have unraveled many of the instructions encoded in the human genome. Taking
advantage of significant advances in parallel sequencing and a wide diversity of other protocols, studies are
proposed in this application to resolve gene-environment interactions that determine variations in the
susceptibility to lung cancer. These studies are timely in light of the worsening levels of environmental pollution
in many US cities that contribute to the development of respiratory disease and to disparities across populations.
Of concern is the persistence of polycyclic aromatic hydrocarbon (PAH) contaminants, such as benzo(a)pyrene
(BaP), and their ability to damage the DNA of lung epithelial cells. While the mutagenicity of BaP has been
extensively characterized, questions remain about the degree to which DNA damage intersects with epigenetic
disruption in driving the overwhelming carcinogenic response of BaP. Genome-wide assessments of BaP toxicity
in the Ramos Laboratory revealed that the parent hydrocarbon and its metabolites activate LINE-1
retrotransposons in lung epithelial cells via epigenetic mechanisms. LINE-1 (Long Interspersed Element-1)
activation is associated with oncogenic signaling, changes in chromatin structure, disruption of epigenetic
control, and malignant transformation. LINE-1 is a family of mobile elements with the ability to copy their own
DNA and use these complementary sequences to randomly insert at other locations within the genome. This
process can be highly mutagenic and associated with genomic instability. We posit that alterations in the lifecycle
of LINE-1 retroelements is a key missing link in our understanding of the complex genetic and epigenetic deficits
associated with BaP carcinogenicity. Of ~100 full-length LINE-1 elements that remain competent for
retrotransposition, nine are recognized as “hot” LINE-1s responsible for the vast majority of pathogenic events.
These hot elements are polymorphic in humans and may therefore contribute to heterogeneity in genetic
susceptibility to environmental PAH toxicity. In response to RFA-ES-20-018, we propose to use in vitro functional
genomics for discovery and validation to test the hypothesis that polymorphic LINE-1 elements differentially
regulate oncogenic signaling in lung epithelial cells and account for differences in susceptibility to BaP. In Aim
1, we will examine hot LINE-1 elements and their constitutive and inducible expression in normal lung and
cancerous epithelial cells by targeted DNA sequencing and single cell RNA sequencing (scRNA-seq). In Aim 2,
we will use CRISPR/dCas9m to edit the LINE-1 promoter to either enhance or silence retrotransposition and its
impact on chromatin structure and transcriptomic landscapes. In Aim 3, we will use computational approaches
to study genetic relationships across variable networks defined by polymorphic LINE-1 variants, with a focus on
TP53, AHR, and RB. These studies will reveal important mechanistic insights into the activation of LINE-1 by
PAH carcinogens and illuminate our understanding of the human variability in carcinogen susceptibility. This
knowledge will lead to better biomarker designs and novel therapies to combat environmental toxicities.
Terms: <3,4-Benzopyrene><3,4-Benzpyrene><Airway health><Antioncogene Protein p53><Aromatic Polycyclic Hydrocarbons><Assay><Automobile Driving><Benzo(a)pyrene><Binding><Bioassay><Biological><Biological Assay><Biological Markers><C-K-RAS><CRISPR><CRISPR editing screen><CRISPR screen><CRISPR-based screen><CRISPR/Cas system><CRISPR/Cas9 screen><Cancer Causing Agents><Cancer cell line><Cancerous><Carcinogen-DNA Adducts><Carcinogens><Cell Body><Cell Communication and Signaling><Cell Line><Cell Respiration><Cell Signaling><CellLine><Cells><Cellular Respiration><Cellular Tumor Antigen P53><Chromatin Structure><Cities><Clustered Regularly Interspaced Short Palindromic Repeats><Complex><DNA><DNA Adducts><DNA Damage><DNA Injury><DNA Methylation><DNA Recombination><DNA seq><DNA sequencing><DNAseq><Deoxyribonucleic Acid><Development><Disease><Disorder><Disparities><Disparity><Elements><Environment><Environmental Carcinogens><Environmental Pollution><Epigenetic><Epigenetic Change><Epigenetic Mechanism><Epigenetic Process><Epithelial Cells><Event><Exposure to><Family><Female><Gene x Environment Interaction><Genes><Genetic><Genetic Polymorphism><Genetic Predisposition><Genetic Predisposition to Disease><Genetic Recombination><Genetic Susceptibility><Genetic propensity><Genetic study><Genome><Genome Instability><Genomic Instability><GxE interaction><Heterogeneity><Human><Human Genome><Hydrocarbons><In Vitro><Inherited Predisposition><Inherited Susceptibility><Instruction><Intracellular Communication and Signaling><Investigation><K-RAS2A><K-RAS2B><K-Ras><K-Ras 2A><K-Ras-2 Oncogene><KRAS><KRAS2><KRAS2 gene><Ki-RAS><Knowledge><Laboratories><Length><Life Cycle><Life Cycle Stages><Light><Link><Liver><Location><Long Interspersed Elements><Lung><Lung Respiratory System><Malignant><Malignant - descriptor><Malignant Tumor of the Lung><Malignant neoplasm of lung><Mediating><Mediator><Mice><Mice Mammals><Modern Man><Molecular Interaction><Murine><Mus><Mutate><NSCLC><NSCLC - Non-Small Cell Lung Cancer><Non-Small Cell Lung Cancer><Non-Small-Cell Lung Carcinoma><Oncogene K-Ras><Oncogenic><Oncogens><Oncoprotein p53><Outcome><P53><Parents><Pathogenicity><Phosphoprotein P53><Phosphoprotein pp53><Photoradiation><Population><Predisposition><Process><Protein TP53><Protocol><Protocols documentation><Pulmonary Cancer><Pulmonary malignant Neoplasm><RASK2><Recombination><Respiratory Disease><Respiratory System Disease><Respiratory System Disorder><Retroelements><Retrotransposition><Retrotransposon><Role><Signal Transduction><Signal Transduction Systems><Signaling><Strains Cell Lines><Susceptibility><TP53><TP53 gene><TRP53><Technology><Testing><Toxic effect><Toxicities><Tumor Protein p53><Tumor Protein p53 Gene><Validation><Variant><Variation><aerobic metabolism><aerobic respiration><bio-markers><biologic><biologic marker><biological signal transduction><biomarker><cancer type><carcinogenicity><clustered regularly interspaced short palindromic repeats screen><combat><constitutive expression><constitutive gene expression><cultured cell line><design><designing><developmental><driving><environment effect on gene><environmental contamination><epigenetically><exposed human population><functional genomics><gene environment interaction><gene manipulation><genetic etiology><genetic manipulation><genetic mechanism of disease><genetic vulnerability><genetically manipulate><genetically perturb><genetically predisposed><genome scale><genome-wide><genomewide><hepatic body system><hepatic organ system><human exposure><human whole genome><in silico><in vivo><inducible expression><inducible gene expression><insight><life course><lung cancer><male><member><mouse model><murine model><new drug treatments><new drugs><new pharmacological therapeutic><new therapeutics><new therapy><next generation therapeutics><novel drug treatments><novel drugs><novel pharmaco-therapeutic><novel pharmacological therapeutic><novel therapeutics><novel therapy><oncogenic agent><oxidative metabolism><p53 Antigen><p53 Genes><p53 Tumor Suppressor><parent><polyaromatic hydrocarbons><polymorphism><polynuclear aromatic hydrocarbon><promoter><promotor><protein p53><pulmonary><renal epithelium><respiratory health><response><scRNA-seq><single cell RNA-seq><single cell RNAseq><single cell expression profiling><single cell transcriptomic profiling><single-cell RNA sequencing><social role><transcriptomics><tumor><v-Ki-RAS2 Kirsten Rat Sarcoma 2 Viral Oncogene Homolog><validations>