Publications by authors named "Howard Cedar"

55 Publications

Chromosomal coordination and differential structure of asynchronous replicating regions.

Nat Commun 2021 02 15;12(1):1035. Epub 2021 Feb 15.

Department of Microbiology and Molecular Genetics, IMRIC, Hebrew University Medical School, Jerusalem, Israel.

Stochastic asynchronous replication timing (AS-RT) is a phenomenon in which the time of replication of each allele is different, and the identity of the early allele varies between cells. By taking advantage of stable clonal pre-B cell populations derived from C57BL6/Castaneous mice, we have mapped the genome-wide AS-RT loci, independently of genetic differences. These regions are characterized by differential chromatin accessibility, mono-allelic expression and include new gene families involved in specifying cell identity. By combining population level mapping with single cell FISH, our data reveal the existence of a novel regulatory program that coordinates a fixed relationship between AS-RT regions on any given chromosome, with some loci set to replicate in a parallel and others set in the anti-parallel orientation. Our results show that AS-RT is a highly regulated epigenetic mark established during early embryogenesis that may be used for facilitating the programming of mono-allelic choice throughout development.
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http://dx.doi.org/10.1038/s41467-021-21348-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7884787PMC
February 2021

Determining gestational age using genome methylation profile: A novel approach for fetal medicine.

Prenat Diagn 2019 10 12;39(11):1005-1010. Epub 2019 Aug 12.

Department of Developmental Biology and Cancer Research, Institute for Medical Research Israel-Canada, Hebrew University Medical School, Jerusalem, Israel.

Gestational age determination by traditional tools (last menstrual period, ultrasonography measurements and Ballard Maturational Assessment in newborns) has major limitations and therefore there is a need to find different approaches. In this study, we looked for a molecular marker that can be used to determine the accurate gestational age of the newborn. To this end, we performed reduced representation bisulfite sequencing (RRBS) on 41 cord blood and matching placenta samples from women between 25 and 40 weeks of gestation and generated an epigenetic clock based on the methylation level at different loci in the genome. We identified a set of 332 differentially methylated regions (DMRs) that undergo demethylation in late gestational age in cord blood cells and can predict the gestational age (r = -.7, P = 2E-05). Once the set of 411 DMRs that undergo de novo methylation in late gestational age was used in combination with the first set, it generated a more accurate clock (R = .77, P = 1.87E-05). We have compared gestational age determined by Ballard score assessment with our epigenetic clock and found high concordance. Taken together, this study demonstrates that DNA methylation can accurately predict gestational age and thus may serve as a good clinical predictor.
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http://dx.doi.org/10.1002/pd.5535DOI Listing
October 2019

Role of transcription complexes in the formation of the basal methylation pattern in early development.

Proc Natl Acad Sci U S A 2018 10 26;115(41):10387-10391. Epub 2018 Sep 26.

Department of Developmental Biology and Cancer Research, Faculty of Medicine, Hebrew University of Jerusalem, 91120 Jerusalem, Israel;

Following erasure in the blastocyst, the entire genome undergoes de novo methylation at the time of implantation, with CpG islands being protected from this process. This bimodal pattern is then preserved throughout development and the lifetime of the organism. Using mouse embryonic stem cells as a model system, we demonstrate that the binding of an RNA polymerase complex on DNA before de novo methylation is predictive of it being protected from this modification, and tethering experiments demonstrate that the presence of this complex is, in fact, sufficient to prevent methylation at these sites. This protection is most likely mediated by the recruitment of enzyme complexes that methylate histone H3K4 over a local region and, in this way, prevent access to the de novo methylation complex. The topological pattern of H3K4me3 that is formed while the DNA is as yet unmethylated provides a strikingly accurate template for modeling the genome-wide basal methylation pattern of the organism. These results have far-reaching consequences for understanding the relationship between RNA transcription and DNA methylation.
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http://dx.doi.org/10.1073/pnas.1804755115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6187119PMC
October 2018

Principles of DNA methylation and their implications for biology and medicine.

Lancet 2018 09 9;392(10149):777-786. Epub 2018 Aug 9.

Department of Developmental Biology and Cancer Research, Hebrew University of Jerusalem, Faculty of Medicine, Jerusalem, Israel. Electronic address:

DNA methylation represents an annotation system for marking the genetic text, thus providing instruction as to how and when to read the information and control transcription. Unlike sequence information, which is inherited, methylation patterns are established in a programmed process that continues throughout development, thus setting up stable gene expression profiles. This DNA methylation paradigm is a key player in medicine. Some changes in methylation closely correlate with age providing a marker for biological ageing, and these same sites could also play a part in cancer. The genome continues to undergo programmed variation in methylation after birth in response to environmental inputs, serving as a memory device that could affect ageing and predisposition to various metabolic, autoimmune, and neurological diseases. Taking advantage of tissue-specific differences, methylation can be used to detect cell death and thereby monitor many common diseases with a simple cell-free circulating-DNA blood test.
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http://dx.doi.org/10.1016/S0140-6736(18)31268-6DOI Listing
September 2018

Postnatal DNA demethylation and its role in tissue maturation.

Nat Commun 2018 05 23;9(1):2040. Epub 2018 May 23.

Department of Developmental Biology and Cancer Research, Hebrew University Medical School, P.O.B. 12272, , Jerusalem, 91120, Israel.

Development in mammals is accompanied by specific de novo and demethylation events that are thought to stabilize differentiated cell phenotypes. We demonstrate that a large percentage of the tissue-specific methylation pattern is generated postnatally. Demethylation in the liver is observed in thousands of enhancer-like sequences associated with genes that undergo activation during the first few weeks of life. Using. conditional gene ablation strategy we show that the removal of these methyl groups is stable and necessary for assuring proper hepatocyte gene expression and function through its effect on chromatin accessibility. These postnatal changes in methylation come about through exposure to hormone signaling. These results define the molecular rules of 5-methyl-cytosine regulation as an epigenetic mechanism underlying cellular responses to. changing environment.
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http://dx.doi.org/10.1038/s41467-018-04456-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5966414PMC
May 2018

Islet cells share promoter hypomethylation independently of expression, but exhibit cell-type-specific methylation in enhancers.

Proc Natl Acad Sci U S A 2017 12 4;114(51):13525-13530. Epub 2017 Dec 4.

Department of Developmental Biology and Cancer Research, Institute For Medical Research Israel-Canada, The Hebrew University-Hadassah Medical School, Jerusalem 91120, Israel;

DNA methylation at promoters is an important determinant of gene expression. Earlier studies suggested that the insulin gene promoter is uniquely unmethylated in insulin-expressing pancreatic β-cells, providing a classic example of this paradigm. Here we show that islet cells expressing insulin, glucagon, or somatostatin share a lack of methylation at the promoters of the insulin and glucagon genes. This is achieved by rapid demethylation of the insulin and glucagon gene promoters during differentiation of Neurogenin3 embryonic endocrine progenitors, regardless of the specific endocrine cell-type chosen. Similar methylation dynamics were observed in transgenic mice containing a human insulin promoter fragment, pointing to the responsible cis element. Whole-methylome comparison of human α- and β-cells revealed generality of the findings: genes active in one cell type and silent in the other tend to share demethylated promoters, while methylation differences between α- and β-cells are concentrated in enhancers. These findings suggest an epigenetic basis for the observed plastic identity of islet cell types, and have implications for β-cell reprogramming in diabetes and diagnosis of β-cell death using methylation patterns of circulating DNA.
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http://dx.doi.org/10.1073/pnas.1713736114DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5754795PMC
December 2017

Programming asynchronous replication in stem cells.

Nat Struct Mol Biol 2017 Dec 13;24(12):1132-1138. Epub 2017 Nov 13.

Department of Developmental Biology and Cancer Research, Hebrew University Medical School, Jerusalem, Israel.

Many regions of the genome replicate asynchronously and are expressed monoallelically. It is thought that asynchronous replication may be involved in choosing one allele over the other, but little is known about how these patterns are established during development. We show that, unlike somatic cells, which replicate in a clonal manner, embryonic and adult stem cells are programmed to undergo switching, such that daughter cells with an early-replicating paternal allele are derived from mother cells that have a late-replicating paternal allele. Furthermore, using ground-state embryonic stem (ES) cells, we demonstrate that in the initial transition to asynchronous replication, it is always the paternal allele that is chosen to replicate early, suggesting that primary allelic choice is directed by preset gametic DNA markers. Taken together, these studies help define a basic general strategy for establishing allelic discrimination and generating allelic diversity throughout the organism.
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http://dx.doi.org/10.1038/nsmb.3503DOI Listing
December 2017

Epigenetic mechanism of FMR1 inactivation in Fragile X syndrome.

Int J Dev Biol 2017 ;61(3-4-5):285-292

Department of Developmental Biology and Cancer Research, Hebrew University, Jerusalem.

Fragile X syndrome is the most frequent cause of inherited intellectual disability. The primary molecular defect in this disease is the expansion of a CGG repeat in the 5' region of the fragile X mental retardation1 (FMR1) gene, leading to de novo methylation of the promoter and inactivation of this otherwise normal gene, but little is known about how these epigenetic changes occur during development. In order to gain insight into the nature of this process, we have used cell fusion technology to recapitulate the events that occur during early embryogenesis. These experiments suggest that the naturally occurring Fragile XFMR1 5' region undergoes inactivation post implantation in a Dicer/Ago-dependent targeted process which involves local SUV39H-mediated tri-methylation of histone H3K9. It thus appears that Fragile X syndrome may come about through inadvertent siRNA-mediated heterochromatinization.
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http://dx.doi.org/10.1387/ijdb.170022hcDOI Listing
June 2018

Annotating the genome by DNA methylation.

Int J Dev Biol 2017 ;61(3-4-5):137-148

Department of Developmental Biology and Cancer Research. Faculty of Medicine, Hebrew University of Jerusalem, Jerusalem, Israel.

DNA methylation plays a prominent role in setting up and stabilizing the molecular design of gene regulation and by understanding this process one gains profound insight into the underlying biology of mammals. In this article, we trace the discoveries that provided the foundations of this field, starting with the mapping of methyl groups in the genome and the experiments that helped clarify how methylation patterns are maintained through cell division. We then address the basic relationship between methyl groups and gene repression, as well as the molecular rules involved in controlling this process during development in vivo. Finally, we describe ongoing work aimed at defining the role of this modification in disease and deciphering how it may serve as a mechanism for sensing the environment.
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http://dx.doi.org/10.1387/ijdb.160270hcDOI Listing
June 2018

Clonally stable Vκ allelic choice instructs Igκ repertoire.

Nat Commun 2017 05 30;8:15575. Epub 2017 May 30.

Department of Developmental Biology and Cancer Research, Institute for Medical Research Israel-Canada, Hebrew University Medical School, Jerusalem 91120, Israel.

Although much has been done to understand how rearrangement of the Igκ locus is regulated during B-cell development, little is known about the way the variable (V) segments themselves are selected. Here we show, using B6/Cast hybrid pre-B-cell clones, that a limited number of V segments on each allele is stochastically activated as characterized by the appearance of non-coding RNA and histone modifications. The activation states are clonally distinct, stable across cell division and developmentally important in directing the Ig repertoire upon differentiation. Using a new approach of allelic ATAC-seq, we demonstrate that the Igκ V alleles have differential chromatin accessibility, which may serve as the underlying basis of clonal maintenance at this locus, as well as other instances of monoallelic expression throughout the genome. These findings highlight a new level of immune system regulation that optimizes gene diversity.
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http://dx.doi.org/10.1038/ncomms15575DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5459994PMC
May 2017

Contribution of epigenetic mechanisms to variation in cancer risk among tissues.

Proc Natl Acad Sci U S A 2017 02 13;114(9):2230-2234. Epub 2017 Feb 13.

Department of Developmental Biology and Cancer Research, Institute for Medical Research Israel-Canada, Faculty of Medicine, The Hebrew University of Jerusalem, Ein Kerem, Jerusalem, Israel 91120;

Recently, it was suggested that tissue variation in cancer risk originates from differences in the number of stem-cell divisions underlying each tissue, leading to different mutation loads. We show that this variation is also correlated with the degree of aberrant CpG island DNA methylation in normal cells. Methylation accumulates during aging in a subset of molecules, suggesting that the epigenetic landscape within a founder-cell population may contribute to tumor formation.
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http://dx.doi.org/10.1073/pnas.1616556114DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5338490PMC
February 2017

DNA Methylation in Cancer and Aging.

Cancer Res 2016 06 2;76(12):3446-50. Epub 2016 Jun 2.

Department of Developmental Biology and Cancer Research, Institute for Medical Research Israel-Canada, Hebrew University Medical School, Jerusalem, Israel.

DNA methylation is known to be abnormal in all forms of cancer, but it is not really understood how this occurs and what is its role in tumorigenesis. In this review, we take a wide view of this problem by analyzing the strategies involved in setting up normal DNA methylation patterns and understanding how this stable epigenetic mark works to prevent gene activation during development. Aberrant DNA methylation in cancer can be generated either prior to or following cell transformation through mutations. Increasing evidence suggests, however, that most methylation changes are generated in a programmed manner and occur in a subpopulation of tissue cells during normal aging, probably predisposing them for tumorigenesis. It is likely that this methylation contributes to the tumor state by inhibiting the plasticity of cell differentiation processes. Cancer Res; 76(12); 3446-50. ©2016 AACR.
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http://dx.doi.org/10.1158/0008-5472.CAN-15-3278DOI Listing
June 2016

Maintenance of Epigenetic Information.

Cold Spring Harb Perspect Biol 2016 05 2;8(5). Epub 2016 May 2.

Department of Developmental Biology and Cancer Research, Institute for Medical Research Israel-Canada, Hebrew University Medical School, Ein Kerem, Jerusalem, Israel 91120.

The genome is subject to a diverse array of epigenetic modifications from DNA methylation to histone posttranslational changes. Many of these marks are somatically stable through cell division. This article focuses on our knowledge of the mechanisms governing the inheritance of epigenetic marks, particularly, repressive ones, when the DNA and chromatin template are duplicated in S phase. This involves the action of histone chaperones, nucleosome-remodeling enzymes, histone and DNA methylation binding proteins, and chromatin-modifying enzymes. Last, the timing of DNA replication is discussed, including the question of whether this constitutes an epigenetic mark that facilitates the propagation of epigenetic marks.
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http://dx.doi.org/10.1101/cshperspect.a019372DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4852805PMC
May 2016

Tissue-specific DNA demethylation is required for proper B-cell differentiation and function.

Proc Natl Acad Sci U S A 2016 May 18;113(18):5018-23. Epub 2016 Apr 18.

Department of Developmental Biology and Cancer Research, Institute for Medical Research Israel-Canada, Hebrew University Medical School, Jerusalem, Israel 91120;

There is ample evidence that somatic cell differentiation during development is accompanied by extensive DNA demethylation of specific sites that vary between cell types. Although the mechanism of this process has not yet been elucidated, it is likely to involve the conversion of 5mC to 5hmC by Tet enzymes. We show that a Tet2/Tet3 conditional knockout at early stages of B-cell development largely prevents lineage-specific programmed demethylation events. This lack of demethylation affects the expression of nearby B-cell lineage genes by impairing enhancer activity, thus causing defects in B-cell differentiation and function. Thus, tissue-specific DNA demethylation appears to be necessary for proper somatic cell development in vivo.
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http://dx.doi.org/10.1073/pnas.1604365113DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4983829PMC
May 2016

Identification of tissue-specific cell death using methylation patterns of circulating DNA.

Proc Natl Acad Sci U S A 2016 Mar 14;113(13):E1826-34. Epub 2016 Mar 14.

Department of Developmental Biology and Cancer Research, Institute for Medical Research Israel-Canada, The Hebrew University-Hadassah Medical School, Jerusalem 91120, Israel;

Minimally invasive detection of cell death could prove an invaluable resource in many physiologic and pathologic situations. Cell-free circulating DNA (cfDNA) released from dying cells is emerging as a diagnostic tool for monitoring cancer dynamics and graft failure. However, existing methods rely on differences in DNA sequences in source tissues, so that cell death cannot be identified in tissues with a normal genome. We developed a method of detecting tissue-specific cell death in humans based on tissue-specific methylation patterns in cfDNA. We interrogated tissue-specific methylome databases to identify cell type-specific DNA methylation signatures and developed a method to detect these signatures in mixed DNA samples. We isolated cfDNA from plasma or serum of donors, treated the cfDNA with bisulfite, PCR-amplified the cfDNA, and sequenced it to quantify cfDNA carrying the methylation markers of the cell type of interest. Pancreatic β-cell DNA was identified in the circulation of patients with recently diagnosed type-1 diabetes and islet-graft recipients; oligodendrocyte DNA was identified in patients with relapsing multiple sclerosis; neuronal/glial DNA was identified in patients after traumatic brain injury or cardiac arrest; and exocrine pancreas DNA was identified in patients with pancreatic cancer or pancreatitis. This proof-of-concept study demonstrates that the tissue origins of cfDNA and thus the rate of death of specific cell types can be determined in humans. The approach can be adapted to identify cfDNA derived from any cell type in the body, offering a minimally invasive window for diagnosing and monitoring a broad spectrum of human pathologies as well as providing a better understanding of normal tissue dynamics.
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http://dx.doi.org/10.1073/pnas.1519286113DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4822610PMC
March 2016

Gender-specific postnatal demethylation and establishment of epigenetic memory.

Genes Dev 2015 May;29(9):923-33

Department of Developmental Biology and Cancer Research, Hebrew University Medical School, Jerusalem 91120, Israel;

DNA methylation patterns are set up in a relatively fixed programmed manner during normal embryonic development and are then stably maintained. Using genome-wide analysis, we discovered a postnatal pathway involving gender-specific demethylation that occurs exclusively in the male liver. This demodification is programmed to take place at tissue-specific enhancer sequences, and our data show that the methylation state at these loci is associated with and appears to play a role in the transcriptional regulation of nearby genes. This process is mediated by the secretion of testosterone at the time of sexual maturity, but the resulting methylation profile is stable and therefore can serve as an epigenetic memory even in the absence of this inducer. These findings add a new dimension to our understanding of the role of DNA methylation in vivo and provide the foundations for deciphering how environment can impact on the epigenetic regulation of genes in general.
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http://dx.doi.org/10.1101/gad.259309.115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4421981PMC
May 2015

Chronic liver inflammation modifies DNA methylation at the precancerous stage of murine hepatocarcinogenesis.

Oncotarget 2015 May;6(13):11047-60

The Goldyne Savad Institute of Gene Therapy, Hadassah-Hebrew University Medical Center, Jerusalem, Israel.

Chronic liver inflammation precedes the majority of hepatocellular carcinomas (HCC). Here, we explore the connection between chronic inflammation and DNA methylation in the liver at the late precancerous stages of HCC development in Mdr2(-/-) (Mdr2/Abcb4-knockout) mice, a model of inflammation-mediated HCC. Using methylated DNA immunoprecipitation followed by hybridization with "CpG islands" (CGIs) microarrays, we found specific CGIs in 76 genes which were hypermethylated in the Mdr2(-/-) liver compared to age-matched healthy controls. The observed hypermethylation resulted mainly from an age-dependent decrease of methylation of the specific CGIs in control livers with no decrease in mutant mice. Chronic inflammation did not change global levels of DNA methylation in Mdr2(-/-) liver, but caused a 2-fold decrease of the global 5-hydroxymethylcytosine level in mutants compared to controls. Liver cell fractionation revealed, that the relative hypermethylation of specific CGIs in Mdr2(-/-) livers affected either hepatocyte, or non-hepatocyte, or both fractions without a correlation between changes of gene methylation and expression. Our findings demonstrate that chronic liver inflammation causes hypermethylation of specific CGIs, which may affect both hepatocytes and non-hepatocyte liver cells. These changes may serve as useful markers of an increased regenerative activity and of a late precancerous stage in the chronically inflamed liver.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4484438PMC
http://dx.doi.org/10.18632/oncotarget.3567DOI Listing
May 2015

A novel pax5-binding regulatory element in the igκ locus.

Front Immunol 2014 23;5:240. Epub 2014 May 23.

Department of Developmental Biology and Cancer Research, Institute for Medical Research Israel-Canada, Hebrew University Medical School , Jerusalem , Israel.

The Igκ locus undergoes a variety of different molecular processes during B cell development, including V(D)J rearrangement and somatic hypermutations (SHM), which are influenced by cis regulatory regions (RRs) within the locus. The Igκ locus includes three characterized RRs termed the intronic (iEκ), 3'Eκ, and Ed enhancers. We had previously noted that a region of DNA upstream of the iEκ and matrix attachment region (MAR) was necessary for demethylation of the locus in cell culture. In this study, we further characterized this region, which we have termed Dm, for demethylation element. Pre-rearranged Igκ transgenes containing a deletion of the entire Dm region, or of a Pax5-binding site within the region, fail to undergo efficient CpG demethylation in mature B cells in vivo. Furthermore, we generated mice with a deletion of the full Dm region at the endogenous Igκ locus. The most prominent phenotype of these mice is reduced SHM in germinal center B cells in Peyer's patches. In conclusion, we propose the Dm element as a novel Pax5-binding cis regulatory element, which works in concert with the known enhancers, and plays a role in Igκ demethylation and SHM.
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http://dx.doi.org/10.3389/fimmu.2014.00240DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4033077PMC
June 2014

Aberrant DNA methylation in ES cells.

PLoS One 2014 22;9(5):e96090. Epub 2014 May 22.

Department of Developmental Biology and Cancer Research, The Hebrew University-Hadassah Medical School, Jerusalem, Israel.

Both mouse and human embryonic stem cells can be differentiated in vitro to produce a variety of somatic cell types. Using a new developmental tracing approach, we show that these cells are subject to massive aberrant CpG island de novo methylation that is exacerbated by differentiation in vitro. Bioinformatics analysis indicates that there are two distinct forms of abnormal de novo methylation, global as opposed to targeted, and in each case the resulting pattern is determined by molecular rules correlated with local pre-existing histone modification profiles. Since much of the abnormal methylation generated in vitro appears to be stably maintained, this modification may inhibit normal differentiation and could predispose to cancer if cells are used for replacement therapy. Excess CpG island methylation is also observed in normal placenta, suggesting that this process may be governed by an inherent program.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0096090PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4031077PMC
June 2015

Molecular rules governing de novo methylation in cancer.

Cancer Res 2014 Mar 22;74(5):1475-83. Epub 2014 Jan 22.

Authors' Affiliations: Department of Developmental Biology and Cancer Research, Institute for Medical Research Israel-Canada, Hebrew University Medical School, Ein Kerem, Jerusalem; Department of Molecular Cell Biology, Weizmann Institute of Science, Rehovot; Department of Computer Sciences, Technion Israel Institute of Technology, Haifa; Agilent Laboratories, Tel Aviv, Israel; and Agilent Technologies, Inc., Santa Clara, California.

De novo methylation of CpG islands is seen in many cancers, but the general rules governing this process are not known. By analyzing DNA from tumors, as well as normal tissues, and by utilizing a range of published data, we have identified a universal set of tumor targets, each with its own "coefficient" of methylation that is largely correlated with its inherent relative ability to recruit polycomb. This pattern is initially formed by a slow process of de novo methylation that occurs during aging and then undergoes expansion early in tumorigenesis, where we hypothesize that it may act as an inhibitor of development-associated gene activation.
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http://dx.doi.org/10.1158/0008-5472.CAN-13-3042DOI Listing
March 2014

Establishment of methylation patterns in ES cells.

Nat Struct Mol Biol 2014 Jan 15;21(1):110-2. Epub 2013 Dec 15.

Department of Developmental Biology and Cancer Research, Institute for Medical Research Israel-Canada, Hebrew University Medical School, Jerusalem, Israel.

After erasure in the early animal embryo, a new bimodal DNA methylation pattern is regenerated at implantation. We have identified a demethylation pathway in mouse embryonic cells that uses hydroxymethylation (Tet1), deamination (Aid), glycosylation (Mbd4) and excision repair (Gadd45a) genes. Surprisingly, this demethylation system is not necessary for generating the overall bimodal methylation pattern but does appear to be involved in resetting methylation patterns during somatic-cell reprogramming.
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http://dx.doi.org/10.1038/nsmb.2734DOI Listing
January 2014

DNA methylation dynamics in health and disease.

Nat Struct Mol Biol 2013 Mar;20(3):274-81

Department of Developmental Biology and Cancer Research, Institute for Medical Research Israel-Canada, Hebrew University Medical School, Jerusalem, Israel.

DNA methylation is an epigenetic mark that is erased in the early embryo and then re-established at the time of implantation. In this Review, dynamics of DNA methylation during normal development in vivo are discussed, starting from fertilization through embryogenesis and postnatal growth, as well as abnormal methylation changes that occur in cancer.
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http://dx.doi.org/10.1038/nsmb.2518DOI Listing
March 2013

Clonal allelic predetermination of immunoglobulin-κ rearrangement.

Nature 2012 Oct 30;490(7421):561-5. Epub 2012 Sep 30.

Department of Developmental Biology and Cancer Research, Institute for Medical Research Israel-Canada, Hebrew University Medical School, POB 12272, Ein Kerem, Jerusalem 91120, Israel.

Although most genes are expressed biallelically, a number of key genomic sites--including immune and olfactory receptor regions--are controlled monoallelically in a stochastic manner, with some cells expressing the maternal allele and others the paternal allele in the target tissue. Very little is known about how this phenomenon is regulated and programmed during development. Here, using mouse immunoglobulin-κ (Igκ) as a model system, we demonstrate that although individual haematopoietic stem cells are characterized by allelic plasticity, early lymphoid lineage cells become committed to the choice of a single allele, and this decision is then stably maintained in a clonal manner that predetermines monoallelic rearrangement in B cells. This is accompanied at the molecular level by underlying allelic changes in asynchronous replication timing patterns at the κ locus. These experiments may serve to define a new concept of stem cell plasticity.
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http://dx.doi.org/10.1038/nature11496DOI Listing
October 2012

Programming of DNA methylation patterns.

Annu Rev Biochem 2012 23;81:97-117. Epub 2012 Feb 23.

Department of Developmental Biology and Cancer Research, Hebrew University Medical School, Ein Kerem, Jerusalem, Israel.

DNA methylation represents a form of genome annotation that mediates gene repression by serving as a maintainable mark that can be used to reconstruct silent chromatin following each round of replication. During development, germline DNA methylation is erased in the blastocyst, and a bimodal pattern is established anew at the time of implantation when the entire genome gets methylated while CpG islands are protected. This brings about global repression and allows housekeeping genes to be expressed in all cells of the body. Postimplantation development is characterized by stage- and tissue-specific changes in methylation that ultimately mold the epigenetic patterns that define each individual cell type. This is directed by sequence information in DNA and represents a secondary event that provides long-term expression stability. Abnormal methylation changes play a role in diseases, such as cancer or fragile X syndrome, and may also occur as a function of aging or as a result of environmental influences.
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http://dx.doi.org/10.1146/annurev-biochem-052610-091920DOI Listing
August 2012

Epigenetics of haematopoietic cell development.

Nat Rev Immunol 2011 Jun 10;11(7):478-88. Epub 2011 Jun 10.

Department of Developmental Biology and Cancer Research, Institute for Medical Research Israel-Canada, Hebrew University Medical School, P.O.B. 12272, Ein Kerem, Jerusalem, Israel, 91120.

Cells of the immune system are generated through a developmental cascade that begins in haematopoietic stem cells. During this process, gene expression patterns are programmed in a series of stages that bring about the restriction of cell potential, ultimately leading to the formation of specialized innate immune cells and mature lymphocytes that express antigen receptors. These events involve the regulation of both gene expression and DNA recombination, mainly through the control of chromatin accessibility. In this Review, we describe the epigenetic changes that mediate this complex differentiation process and try to understand the logic of the programming mechanism.
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http://dx.doi.org/10.1038/nri2991DOI Listing
June 2011

Genome-wide de novo methylation in epithelial ovarian cancer.

Int J Gynecol Cancer 2011 Feb;21(2):269-79

Department of Gynecology, Shaare Zedek Medical Center, Israel.

Background: DNA methylation regulates gene expression during development. The methylation pattern is established at the time of implantation. CpG islands are genome regions usually protected from methylation; however, selected islands are methylated later. Many undergo methylation in cancer, causing epigenetic gene silencing. Aberrant methylation occurs early in tumorigenesis, in a specific pattern, inhibiting differentiation.Although methylation of specific genes in ovarian tumors has been demonstrated in numerous studies, they represent only a fraction of all methylated genes in tumorigenesis.

Objectives: To explore the hypermethylation design in ovarian cancer compared with the methylation profile of normal ovaries, on a genome-wide scale, thus shedding light on the role of gene silencing in ovarian carcinogenesis.Identifying genes that undergo de novo methylation in ovarian cancer may assist in creating biomarkers for disease diagnosis, prognosis, and treatment responsiveness.

Methods: DNA was collected from human epithelial ovarian cancers and normal ovaries. Methylation was detected by immunoprecipitation using 5-methyl-cytosine-antibodies. DNA was hybridized to a CpG island microarray containing 237,220 gene promoter probes. Results were analyzed by hybridization intensity, validated by bisulfite analysis.

Results: : A total of 367 CpG islands were specifically methylated in cancer cells. There was enrichment of methylated genes in functional categories related to cell differentiation and proliferation inhibition. It seems that their silencing enables tumor proliferation.

Conclusions: This study provides new perspectives on methylation in ovarian carcinoma, genome-wide. It illustrates how methylation of CpG islands causes silencing of genes that have a role in cell differentiation and functioning. It creates potential biomarkers for diagnosis, prognosis, and treatment responsiveness.
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http://dx.doi.org/10.1097/IGC.0b013e31820e5cdaDOI Listing
February 2011

Epigenetic control of recombination in the immune system.

Semin Immunol 2010 Dec 15;22(6):323-9. Epub 2010 Sep 15.

Department of Developmental Biology and Cancer Research, The Hebrew University, Hadassah Medical School, Jerusalem 91120, Israel.

Immune receptor gene expression is regulated by a series of developmental events that modify their accessibility in a locus, cell type, stage and allele-specific manner. This is carried out by a programmed combination of many different molecular mechanisms, including region-wide replication timing, changes in nuclear localization, chromatin contraction, histone modification, nucleosome positioning and DNA methylation. These modalities ultimately work by controlling steric interactions between receptor loci and the recombination machinery.
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http://dx.doi.org/10.1016/j.smim.2010.07.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2981633PMC
December 2010

Reprogramming of DNA replication timing.

Stem Cells 2010 Mar;28(3):443-9

The Hadassah Human Embryonic Stem Cells Research Center, Goldyne-Savad Institute of Gene Therapy, Department of OB & GYN, Hadassah University Hospital, Jerusalem 91120, Israel.

Replication timing is an important developmentally regulated regional property that is correlated with chromosome structure and gene expression, but little is known about the establishment and maintenance of these patterns. Here we followed the fate of replication timing patterns in cells that undergo reprogramming either through somatic-cell nuclear transplantation or by the generation of induced pluripotential stem cells. We have investigated three different paradigms, stage-specific replication timing, parental allele-specific asynchrony (imprinted regions), and random allelic asynchronous replication. In all cases, somatic replication timing patterns were reset exactly at the appropriate stage in early development and could be properly established upon re-differentiation. Taken together, these results suggest that, unlike DNA methylation, the molecular mechanisms governing replication timing are not only stable but can also be easily reprogrammed.
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http://dx.doi.org/10.1002/stem.303DOI Listing
March 2010

Allelic inactivation of rDNA loci.

Genes Dev 2009 Oct;23(20):2437-47

Department of Cellular Biochemistry and Experimental Medicine, Hebrew University Medical School, Ein Kerem, Jerusalem 91120, Israel.

Human cells contain several hundred ribosomal genes (rDNA) that are clustered into nucleolar organizer regions (NORs) on the short arms of five different acrocentric chromosomes. Only approximately 50% of the gene copies are actually expressed in somatic cells. Here, we used a new cytological technique to demonstrate that rDNA is regulated allelically in a regional manner, with one parental copy of each NOR being repressed in any individual cell. This process is similar to that of X-chromosome inactivation in females. Early in development, one copy of each NOR becomes late-replicating, thus probably marking it for inactivation and subsequent targeted de novo methylation at rDNA promoter regions. Once established, this multichromosomal allelic pattern is then maintained clonally in somatic cells. This pathway may serve as an epigenetic mechanism for controlling the number of available rDNA copies during development.
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http://dx.doi.org/10.1101/gad.544509DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2764490PMC
October 2009