Histone modification and cytosine-5 DNA methylation represent two importantepigenetic marks. These two very distinct modifications work in close cooperation to controltranscriptional potential, thereby influencing a wide diversity of biological processes. The pastdecade has seen an explosion of interest in the role of these epigenetic modifications inhuman heath and disease. The goal of the proposed application is to advance ourunderstanding of these two modifications, by developing and applying two novel in vivoimaging technologies capable of visualizing the epigenetic and even genetic consequences ofthese modifications. The first specific aim is to develop an in vivo imaging system to assess the causalinfluence of individual epigenetic modifications. The technology is designed to both achieveselective introduction of a desired modification at a predetermined genomic locus, and toproduce a quantitative optical readout of the effect of the epigenetic modification ontranscription. This is achieved through the sequence-specific recruitment of epigenetic modifierproteins to a promoter driving a fluorescence-luminescence fusion reporter. The power of thisapproach comes from the combination of the quantitative optical analysis and the localizedrecruitment. This will allow high-throughput evaluation of the primary causal effects ofindividual epigenetic modifications. The second specific aim is to develop an imaging assay for the in vivo visualization ofCpG transition mutations. Transition mutations at the epigenetic DNA methylation mark areresponsible for approximately one-third of all human hereditary disease mutations and fornearly 50% of all p53 point mutations found in human colorectal cancer. However, the lack ofimaging tools for this type of epigenetically induced mutagenesis has held back ourunderstanding of the timing and cell-type specificity of this event in vivo. The proposed systemis designed to provide direct visualization of the result of the mutation event. This is achievedthrough the use of a mutant green fluorescent protein that produces a fluorescent signal uponCpG transition mutation in the chromophore region. The conversion of a specific geneticmutation into an optical signal provides an attractive opportunity to analyze the mutationwithout employing direct sequencing. The in situ analysis of mutagenesis would allow us to notonly determine tissue-specific and cell-type specific in vivo CpG mutation frequencies, but alsoto analyze mutation kinetics.
During the past decade; there has been an increasing recognition of epigeneticcontribution to pathogenesis. This study aims to develop imaging technologies to advance ourunderstanding of the causal role of selected epigenetic marks on transcriptional potential andon mutagenesis in vivo; with the long term goal of using this knowledge to improve ourtherapeutic or diagnostic strategies targeting epigenetic modifications.
| Lee, Kwang-Ho; Oghamian, Shirley; Park, Jin-A et al. (2017) The REMOTE-control system: a system for reversible and tunable control of endogenous gene expression in mice. Nucleic Acids Res 45:12256-12269 |
| Oh, Edward; Jeremian, Richie; Oh, Gabriel et al. (2017) Transcriptional heterogeneity in the lactase gene within cell-type is linked to the epigenome. Sci Rep 7:41843 |
| Lee, Kwang-Ho; Laird, Peter W (2016) All Things in Moderation: Prevention of Intestinal Adenomas by DNA Hypomethylation. Cancer Prev Res (Phila) 9:509-11 |
| Shen, Hui; Laird, Peter W (2013) Interplay between the cancer genome and epigenome. Cell 153:38-55 |
