Taberlay P.C.,Garvan Institute of Medical Research |
Taberlay P.C.,University of Southern California |
Taberlay P.C.,University of New South Wales |
Statham A.L.,Garvan Institute of Medical Research |
And 6 more authors.
Genome Research | Year: 2014
It is well established that cancer-associated epigenetic repression occurs concomitant with CpG island hypermethylation and loss of nucleosomes at promoters, but the role of nucleosome occupancy and epigenetic reprogramming at distal regulatory elements in cancer is still poorly understood. Here, we evaluate the scope of global epigenetic alterations at enhancers and insulator elements in prostate and breast cancer cells using simultaneous genome-wide mapping of DNA methylation and nucleosome occupancy (NOMe-seq). We find that the genomic location of nucleosome-depleted regions (NDRs) is mostly cell type specific and preferentially found at enhancers in normal cells. In cancer cells, however, we observe a global reconfiguration of NDRs at distal regulatory elements coupled with a substantial reorganization of the cancer methylome. Aberrant acquisition of nucleosomes at enhancer-associated NDRs is associated with hypermethylation and epigenetic silencing marks, and conversely, loss of nucleosomes with demethylation and epigenetic activation. Remarkably, we show that nucleosomes remain strongly organized and phased at many facultative distal regulatory elements, even in the absence of a NDR as an anchor. Finally, we find that key transcription factor (TF) binding sites also show extensive peripheral nucleosome phasing, suggesting the potential for TFs to organize NDRs genome-wide and contribute to deregulation of cancer epigenomes. Together, our findings suggest that ''decommissioning'' of NDRs and TFs at distal regulatory elements in cancer cells is accompanied by DNA hypermethylation susceptibility of enhancers and insulator elements, which in turn may contribute to an altered genome-wide architecture and epigenetic deregulation in malignancy. © 2014 Totoki et al.; Published by Cold Spring Harbor Laboratory Press. Source
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 224.83K | Year: 2014
DESCRIPTION (provided by applicant): Epigenetic mechanisms regulate gene expression potential and alterations in epigenetic marks, such as DNA methylation and histone modifications, have been associated with a variety of diseases. Tissue samples (both research and clinical) are often preserved by formalin fixation and paraffin embedment (FFPE), which allows for long term storage in minimally controlled environments. However there can be significant variability in the preparation of FFPE samples and the preservation conditions are harsh, making FFPE samples challenging for sophisticated downstream analyses. Clinical FFPE samples are often accompanied by valuable information including, histology, treatment course and patient outcome, thus they are an incredibly valuable source for linking basic and clinical research. The goal of this Phase I proposal is to provide the feasibility studies to use or newly developed transposase associate chromatin immunoprecipitation (TA-ChIP) to localize DNA methylation and
Agency: NSF | Branch: Standard Grant | Program: | Phase: | Award Amount: 150.00K | Year: 2012
This Small Business Innovation Research (SBIR) Phase I project proposes to develop a high-throughput, multi-analyte chromatin immunoprecipitation (ChIP) system. ChIP is a widely used technique among life science and biomedical researchers seeking map the specific DNA sequences that are bound by a DNA binding protein. Its use ranges from single gene analysis through to genome-wide applications that use next generation sequencing (NGS) technologies as readout. As envisioned, the proposed technology will transform the lengthy and cumbersome multi-day ChIP into a high throughput compatible single-day experiment. The technology will result in the insertion of oligonucleotides containing NGS platform-compatible tags and bar-code sequences into DNA fragments associated with the immunoprecipitating antibodies. Each antibody will be associated with a unique barcode that can also be used for direct DNA sequencing or PCR analysis. Feasibility of this approach will be achieved by systematic identification of optimal conditions for insertion of the bar-code containing oligonucleotides into chromatin, and then, with antibody-coupled oligonucleotides. Final validation will be performed at the genome scale using NGS to compare the genomic representation of the library produced by the novel method with that of traditional ChIP.
The broader impact/commercial potential of this project is far reaching. The life sciences research tools market is currently estimated at $42 billion, with the epigenetic sector experiencing high growth fueled by researchers purchasing commercial epigenetic products rather than spending the time and effort to develop them in-house, and by advances in NGS, which has accelerated genome-wide epigenetic analyses. Successful development of the high-throughput, multi-analyte ChIP will have significant impact scientifically and commercially in the life sciences and biomedical research arenas. In less than five years post-launch, it is projected to replace traditional ChIP, which represents 20-25% of the epigenetic research tools market estimated at $175-245M in 2010. The development of this method will open epigenetic analysis to virtually all researchers by eliminating technical barriers, and by significantly reducing sample size requirements associated with traditional ChIP, including a potential for single cell analysis. Development of this technology will spur the creation of additional novel technologies such as homogeneous ChIP for high throughput screening, multi-analyte ChIP, and open the door for environmental, nutrition, and toxicology disciplines to study the epigenetic profiles of any eukaryotic organism on a genome wide scale.
Active Motif, Inc. | Date: 2014-05-22
Disclosed herein are compositions, methods and kits useful for epigenetic analysis based on the use of transposons that are targeted to specific regions of chromatin based on DNA-DNA interactions, protein-protein interactions, RNA-RNA interactions, and nucleic acid-protein interactions.
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 314.07K | Year: 2010
DESCRIPTION (provided by applicant): Depression is the leading cause of disability among individuals between the ages of 15-44, affecting approximately 21 million Americans and is the leading cause of suicide. Current diagnostic tools available for medical professionals are limited to questionnaires and other inherently subjective approaches. Improving accuracy of diagnosis has been cited as a pressing need in the field. Recent advances have shed light on the genetic basis for psychopathological conditions, which not only includes changes in gene expression, but also epigenomic changes as well. Epigenomic mechanisms, which regulate gene activity without altering the DNA code, consist fundamentally of DNA methylation of CpG-dinucleotides, which occurs at the fifth position of the cytosine pyrimidine ring, and regulation of chromatin structure through post-translational modification of histones. In this Phase I proposal , we intend to develop a Clinical Chromatin ImmunoPrecipitation assay (C- ChIP), for use with formaldehyde-fixed paraffin embedded (FFPE) human brain specimens, which will enable analysis of changes in histone post-translational modifications in normal and suicide brains samples. Chromatin immunoprecipitation is a powerful technique that captures DNA bound proteins, and enables quantification of the specific immunoprecipitated DNA sequences relative to input. Development of the C- ChIP assay will be achieved by first establishing a rat tissue model system, and in conjunction with the isolation of a panel of highly characterized and specific monoclonal antibodies to histone modifications associated with either transcriptionally active or repressed loci, will be used to systematically adapt existing ChIP protocols developed for in vitro cultured cells, into a highly sensitive assay compatible for use with FFPE sections. Assay validation with clinical samples will be performed collaboratively on a rat maternal care model and subsequently on FFPE control and suicide brain samples. The successful development of the C-ChIP assay will have two effects. First, would be the commercialization of the C-ChIP assay for the broad research market, providing for the first time, an assay which enables the functional genomic analysis of archived clinical samples. The second outcome of this Phase I application is that the C-ChIP assay will enable subsequent Phase II studies in which genomic-wide survey of a large cohort of suicide brains specimens will be examined for epigenetic alternations which could possibly serve as biomarkers for depression. With this information, it may be possible to subsequently identify peripheral markers that correlate with the brain markers, leading to the development of a diagnostic assay for assessing depression and suicide risk. PUBLIC HEALTH RELEVANCE: 1-3 Suicide and depression are major public health concerns . Recent studies have identified some of the molecular mechanisms involved in suicide and depression and these 7-15 mechanisms include changes in the regulation of gene expression in the brain . This Phase 1 application describes the development of an assay that will enable identification of gene expression changes through the whole genome, which could subsequently lead to the development of a much needed diagnostic for depression and suicide risk.