Epigenetic regulation of programmed genome instability in O. trifallax

Summary

Principal Investigator: JOHN RUSSELL BRACHT
Abstract: DESCRIPTION (provided by applicant): Cytosine methylation of DNA is a repressive chromatin mark important for silencing genes during mammalian development, and cancer cells frequently silence tumor suppressor genes by aberrantly methylating their promoters. Genome stability is compromised when DNA methylation patterns are disrupted in cancer or in developmental disease. Despite the critical role of DNA methylation in gene expression and genome stability, the mechanisms guiding de novo DNA methylation remain poorly defined. Oxytricha trifallax is a unicellular eukaryote that performs dramatic genome rearrangements in a developmental process that transforms its micronucleus (MIC) genome into a differentiated macronucleus (MAC). This process is accomplished by the precise elimination of 95% of the genome, and recapitulates the key property of stem cells: self-renewal (one copy of the MIC is retained at all times) and differentiation (MAC formation). Interestingly, my work in the Landweber lab has shown that this organism uses de novo DNA methylation in the elimination process. The fact that most of the Oxytricha genome is methylated and eliminated makes it an attractive model system to understand how methylation is targeted: aberrant methylation and subsequent elimination of the 5% retained DNA would be fatal. In addition, the reproducibility of genome rearrangements in this ciliate makes it a unique model for studying the connection between DNA methylation and genome stability, both of which are relevant to cancer and developmental processes generally. AIM 1: Test the hypothesis that methylation induces DNA degradation in Oxytricha through microinjection of artificially constructed, in vitro methylated chromosomes into the MAC of vegetative cells. AIM 2: Methyl-DNA immunoprecipitation and high throughput sequencing (meDIP-Seq) will be used to define the endogenous domains of de novo DNA methylation during genome rearrangements in Oxytricha. AIM 3: The functional relevance of cytosine methylation in Oxytricha genome rearrangements will be analyzed by use of chemical inhibitors of methyltransferases, followed by deep sequencing the DNA from treated cells. AIM 4: Identification of novel methylation pathway proteins by methyl-cytosine chromatin IP (me-ChIP) and mass-spec analysis. Oxytricha trifallax provides an unprecedented opportunity to study the role of DNA methylation in a model system that has evolved an elaborate genome rearrangement process. Currently, the targeting of de novo methylation in human disease is poorly understood, so insights from Oxytricha will be immediately relevant to studies of human health and disease. Any conserved candidate proteins or motifs identified in the ciliate model can be tested in human tissue culture or mouse cancer models for potential oncogenic roles.
Funding Period: 2012-08-01 - 2014-07-31
more information: NIH RePORT

Top Publications

  1. pmc Piwi-interacting RNAs protect DNA against loss during Oxytricha genome rearrangement
    Wenwen Fang
    Department of Molecular Biology, Princeton University, Princeton, NJ 08544, USA
    Cell 151:1243-55. 2012
  2. pmc Genomes on the edge: programmed genome instability in ciliates
    John R Bracht
    Department of Ecology and Evolutionary Biology, Princeton University, Princeton, NJ 08544, USA
    Cell 152:406-16. 2013
  3. pmc The Oxytricha trifallax macronuclear genome: a complex eukaryotic genome with 16,000 tiny chromosomes
    Estienne C Swart
    Department of Ecology and Evolutionary Biology, Princeton University, Princeton, New Jersey, United States of America
    PLoS Biol 11:e1001473. 2013
  4. pmc Beyond transcriptional silencing: is methylcytosine a widely conserved eukaryotic DNA elimination mechanism?
    John R Bracht
    Department of Ecology and Evolutionary Biology, Princeton University, Guyot Hall, Princeton, NJ, USA
    Bioessays 36:346-52. 2014

Research Grants

Detail Information

Publications5

  1. pmc Piwi-interacting RNAs protect DNA against loss during Oxytricha genome rearrangement
    Wenwen Fang
    Department of Molecular Biology, Princeton University, Princeton, NJ 08544, USA
    Cell 151:1243-55. 2012
    ..Our findings highlight small RNAs as powerful transgenerational carriers of epigenetic information for genome programming...
  2. pmc Genomes on the edge: programmed genome instability in ciliates
    John R Bracht
    Department of Ecology and Evolutionary Biology, Princeton University, Princeton, NJ 08544, USA
    Cell 152:406-16. 2013
    ....
  3. pmc The Oxytricha trifallax macronuclear genome: a complex eukaryotic genome with 16,000 tiny chromosomes
    Estienne C Swart
    Department of Ecology and Evolutionary Biology, Princeton University, Princeton, New Jersey, United States of America
    PLoS Biol 11:e1001473. 2013
    ..trifallax macronuclear genome will provide an invaluable resource for investigating programmed genome rearrangements, complementing studies of rearrangements arising during evolution and disease...
  4. pmc Beyond transcriptional silencing: is methylcytosine a widely conserved eukaryotic DNA elimination mechanism?
    John R Bracht
    Department of Ecology and Evolutionary Biology, Princeton University, Guyot Hall, Princeton, NJ, USA
    Bioessays 36:346-52. 2014
    ....

Research Grants30

  1. The Shelf Live Evaluation of Investigational Dosage Forms
    Jonathan White; Fiscal Year: 2013
    ..This contract is essential for continued assurance of the quality of drugs undergoing clinical investigation for different types of cancer by Cancer Therapeutics Evaluation Program. ..
  2. University of Maryland Greenebaum Cancer Center Support Grant
    Kevin J Cullen; Fiscal Year: 2013
    ..Reflecting our remarkable and continued growth, UMGCC seeks to renew its CCSG to enhance and expand its efforts in high-quality and clinically relevant cancer research. ..
  3. CENTER FOR GASTROINTESTINAL BIOLOGY AND DISEASE
    Robert S Sandler; Fiscal Year: 2013
    ..Through all of its activities, the Center improves communication, promotes collaboration, develops careers and generally enriches the intellectual climate for digestive disease research. ..
  4. Intellectual and Development Disabilities Research Center
    Marc Yudkoff; Fiscal Year: 2013
    ..5 million from NICHD). The Center includes an excess of 70 Penn faculty at 15 departments at the Schools of Medicine, Veterinary Medicine, Nursing, the Wistar Institute, and the College of Arts and Sciences. ..
  5. NK Cells, Their Receptors and Unrelated Donor Transplant
    Jeffrey S Miller; Fiscal Year: 2013
    ..The program also promises to change practice of allogeneic transplantation, to the greater benefit of patients with advanced leukemia. ..
  6. STATISTICAL METHODS FOR MEDICAL STUDIES
    Ross L Prentice; Fiscal Year: 2013
    ..abstract_text> ..
  7. The MIT Center for Single-Cell Dynamics in Cancer (SCDC)
    Scott R Manalis; Fiscal Year: 2013
    ..These facilities and all reagents generated by the cores will be made available to other PS-OCs. ..
  8. M D Anderson Cancer Center Prosate SPORE
    Christopher J Logothetis; Fiscal Year: 2013
    ..We are optimistic that our research efforts will contribute to reductions in the incidence of, and morbidity and mortality from, this devastating disease by translating basic research into clinical practice. ..
  9. Structural Cell Biology of DNA Repair Machines
    John A Tainer; Fiscal Year: 2013
    ..abstract_text> ..
  10. Longitudinal Epigenomic Tracking of the Reprogramming Process
    Michael P Snyder; Fiscal Year: 2013
    ..By understanding the epigenomic landscapes of different tissues and ages and tracking changes during reprogramming, we hope to identify even more features associated with successful and abortive de-differentiation. ..
  11. Experimental Therapeutics of Leukemia
    John C Byrd; Fiscal Year: 2013
    ..We believe that this SPORE group, as a multidisciplinary, highly interactive and accomplished team, will have a substantial impact on improving the clinical outcome of leukemia patients. ..
  12. Genetics of DNA methylation patterning of arabidopsis.
    Steven E Jacobsen; Fiscal Year: 2013
    ..Thus a further understanding of the DNA methylation may someday lead to methods for correcting DNA methylation patterning defects. ..
  13. Mechanistic Pharmacology of Anti-Mitotics and Apoptosis Regulation
    Timothy J Mitchison; Fiscal Year: 2013
    ..In aim 4 we will pursue several approaches towards translating mechanistic understanding from aims 1-3 into improved patient care. ..
  14. CSHL CANCER CENTER SUPPORT GRANT
    BRUCE W STILLMAN; Fiscal Year: 2013
    ....
  15. UNMC EPPLEY CANCER CENTER SUPPORT GRANT
    Kenneth H Cowan; Fiscal Year: 2013
    ....
  16. Baylor Intellectual and Developmental Disabilities Research Center
    Huda Y Zoghbi; Fiscal Year: 2013
    ..abstract_text> ..
  17. B-cell Biology of Mucosal Immune Protection from SIV Challenge
    Eric Hunter; Fiscal Year: 2013
    ....