Publications by authors named "Mamta Tahiliani"

14 Publications

  • Page 1 of 1

A Susceptibility Locus on Chromosome 13 Profoundly Impacts the Stability of Genomic Imprinting in Mouse Pluripotent Stem Cells.

Cell Rep 2020 03;30(11):3597-3604.e3

Skirball Institute of Biomolecular Medicine, Department of Cell Biology, NYU Langone Medical Center, New York, NY 10016, USA; Helen L. and Martin S. Kimmel Center for Biology and Medicine, NYU Langone Medical Center, New York, NY 10016, USA; Laura and Isaac Perlmutter Cancer Center, NYU Langone Medical Center, New York, NY 10016, USA; Sanford I. Weill Department of Medicine, Sandra and Edward Meyer Cancer Center, Weill Cornell Medicine, New York, NY 10021, USA. Electronic address:

Cultured pluripotent cells accumulate detrimental chromatin alterations, including DNA methylation changes at imprinted genes known as loss of imprinting (LOI). Although the occurrence of LOI is considered a stochastic phenomenon, here we document a genetic determinant that segregates mouse pluripotent cells into stable and unstable cell lines. Unstable lines exhibit hypermethylation at Dlk1-Dio3 and other imprinted loci, in addition to impaired developmental potential. Stimulation of demethylases by ascorbic acid prevents LOI and loss of developmental potential. Susceptibility to LOI greatly differs between commonly used mouse strains, which we use to map a causal region on chromosome 13 with quantitative trait locus (QTL) analysis. Our observations identify a strong genetic determinant of locus-specific chromatin abnormalities in pluripotent cells and provide a non-invasive way to suppress them. This highlights the importance of considering genetics in conjunction with culture conditions for assuring the quality of pluripotent cells for biomedical applications.
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http://dx.doi.org/10.1016/j.celrep.2020.02.073DOI Listing
March 2020

TET methylcytosine oxidases: new insights from a decade of research.

J Biosci 2020 ;45

Division of Signaling and Gene Expression, La Jolla Institute for Immunology, La Jolla, CA 92037, USA.

In mammals, DNA methyltransferases transfer a methyl group from S-adenosylmethionine to the 5 position of cytosine in DNA. The product of this reaction, 5-methylcytosine (5mC), has many roles, particularly in suppressing transposable and repeat elements in DNA. Moreover, in many cellular systems, cell lineage specification is accompanied by DNA demethylation at the promoters of genes expressed at high levels in the differentiated cells. However, since direct cleavage of the C-C bond connecting the methyl group to the 5 position of cytosine is thermodynamically disfavoured, the question of whether DNA methylation was reversible remained unclear for many decades. This puzzle was solved by our discovery of the TET (Ten- Eleven Translocation) family of 5-methylcytosine oxidases, which use reduced iron, molecular oxygen and the tricarboxylic acid cycle metabolite 2-oxoglutarate (also known as a-ketoglutarate) to oxidise the methyl group of 5mC to 5-hydroxymethylcytosine (5hmC) and beyond. TET-generated oxidised methylcytosines are intermediates in at least two pathways of DNA demethylation, which differ in their dependence on DNA replication. In the decade since their discovery, TET enzymes have been shown to have important roles in embryonic development, cell lineage specification, neuronal function and cancer. We review these findings and discuss their implications here.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7216820PMC
October 2020

Overlapping Requirements for Tet2 and Tet3 in Normal Development and Hematopoietic Stem Cell Emergence.

Cell Rep 2015 Aug 6;12(7):1133-43. Epub 2015 Aug 6.

Developmental Biology Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. Electronic address:

The Tet family of methylcytosine dioxygenases (Tet1, Tet2, and Tet3) convert 5-methylcytosine to 5-hydroxymethylcytosine. To date, functional overlap among Tet family members has not been examined systematically in the context of embryonic development. To clarify the potential for overlap among Tet enzymes during development, we mutated the zebrafish orthologs of Tet1, Tet2, and Tet3 and examined single-, double-, and triple-mutant genotypes. Here, we identify Tet2 and Tet3 as the major 5-methylcytosine dioxygenases in the zebrafish embryo and uncover a combined requirement for Tet2 and Tet3 in hematopoietic stem cell (HSC) emergence. We demonstrate that Notch signaling in the hemogenic endothelium is regulated by Tet2/3 prior to HSC emergence and show that restoring expression of the downstream gata2b/scl/runx1 transcriptional network can rescue HSCs in tet2/3 double mutant larvae. Our results reveal essential, overlapping functions for tet genes during embryonic development and uncover a requirement for 5hmC in regulating HSC production.
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http://dx.doi.org/10.1016/j.celrep.2015.07.025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4545447PMC
August 2015

Simultaneous sequencing of oxidized methylcytosines produced by TET/JBP dioxygenases in Coprinopsis cinerea.

Proc Natl Acad Sci U S A 2014 Dec 18;111(48):E5149-58. Epub 2014 Nov 18.

Division of Signaling and Gene Expression, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037; Department of Pharmacology and Moores Cancer Center, University of California, San Diego, La Jolla, CA 92093

TET/JBP enzymes oxidize 5-methylpyrimidines in DNA. In mammals, the oxidized methylcytosines (oxi-mCs) function as epigenetic marks and likely intermediates in DNA demethylation. Here we present a method based on diglucosylation of 5-hydroxymethylcytosine (5hmC) to simultaneously map 5hmC, 5-formylcytosine, and 5-carboxylcytosine at near-base-pair resolution. We have used the method to map the distribution of oxi-mC across the genome of Coprinopsis cinerea, a basidiomycete that encodes 47 TET/JBP paralogs in a previously unidentified class of DNA transposons. Like 5-methylcytosine residues from which they are derived, oxi-mC modifications are enriched at centromeres, TET/JBP transposons, and multicopy paralogous genes that are not expressed, but rarely mark genes whose expression changes between two developmental stages. Our study provides evidence for the emergence of an epigenetic regulatory system through recruitment of selfish elements in a eukaryotic lineage, and describes a method to map all three different species of oxi-mCs simultaneously.
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http://dx.doi.org/10.1073/pnas.1419513111DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4260599PMC
December 2014

Expanding the epigenetic landscape: novel modifications of cytosine in genomic DNA.

Cold Spring Harb Perspect Biol 2014 Oct 1;6(10):a018630. Epub 2014 Oct 1.

Skirball Institute/NYU School of Medicine, New York, New York 10016.

Methylation of the base cytosine in DNA is critical for silencing endogenous retroviruses, regulating gene expression, and establishing cellular identity, and has long been regarded as an indelible epigenetic mark. The recent discovery that the ten eleven translocation (TET) proteins can oxidize 5-methylcytosine (5mC) resulting in the formation of 5-hydroxymethylcytosine (5hmC) and other oxidized cytosine variants in the genome has triggered a paradigm shift in our understanding of how dynamic changes in DNA methylation regulate transcription and cellular differentiation, thus influencing normal development and disease.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4176005PMC
http://dx.doi.org/10.1101/cshperspect.a018630DOI Listing
October 2014

Genome-wide mapping of 5-hydroxymethylcytosine in embryonic stem cells.

Nature 2011 May 8;473(7347):394-7. Epub 2011 May 8.

Harvard Medical School, Immune Disease Institute and Program in Cellular and Molecular Medicine, Children's Hospital Boston, Boston, Massachusetts 02115, USA.

5-hydroxymethylcytosine (5hmC) is a modified base present at low levels in diverse cell types in mammals. 5hmC is generated by the TET family of Fe(II) and 2-oxoglutarate-dependent enzymes through oxidation of 5-methylcytosine (5mC). 5hmC and TET proteins have been implicated in stem cell biology and cancer, but information on the genome-wide distribution of 5hmC is limited. Here we describe two novel and specific approaches to profile the genomic localization of 5hmC. The first approach, termed GLIB (glucosylation, periodate oxidation, biotinylation) uses a combination of enzymatic and chemical steps to isolate DNA fragments containing as few as a single 5hmC. The second approach involves conversion of 5hmC to cytosine 5-methylenesulphonate (CMS) by treatment of genomic DNA with sodium bisulphite, followed by immunoprecipitation of CMS-containing DNA with a specific antiserum to CMS. High-throughput sequencing of 5hmC-containing DNA from mouse embryonic stem (ES) cells showed strong enrichment within exons and near transcriptional start sites. 5hmC was especially enriched at the start sites of genes whose promoters bear dual histone 3 lysine 27 trimethylation (H3K27me3) and histone 3 lysine 4 trimethylation (H3K4me3) marks. Our results indicate that 5hmC has a probable role in transcriptional regulation, and suggest a model in which 5hmC contributes to the 'poised' chromatin signature found at developmentally-regulated genes in ES cells.
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http://dx.doi.org/10.1038/nature10102DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3124347PMC
May 2011

Tet1 and Tet2 regulate 5-hydroxymethylcytosine production and cell lineage specification in mouse embryonic stem cells.

Cell Stem Cell 2011 Feb;8(2):200-13

Immune Disease Institute and Program in Cellular and Molecular Medicine, Children's Hospital Boston, Boston, MA 02115, USA.

TET family enzymes convert 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) in DNA. Here, we show that Tet1 and Tet2 are Oct4-regulated enzymes that together sustain 5hmC in mouse embryonic stem cells (ESCs) and are induced concomitantly with 5hmC during reprogramming of fibroblasts to induced pluripotent stem cells. ESCs depleted of Tet1 by RNAi show diminished expression of the Nodal antagonist Lefty1 and display hyperactive Nodal signaling and skewed differentiation into the endoderm-mesoderm lineage in embryoid bodies in vitro. In Fgf4- and heparin-supplemented culture conditions, Tet1-depleted ESCs activate the trophoblast stem cell lineage determinant Elf5 and can colonize the placenta in midgestation embryo chimeras. Consistent with these findings, Tet1-depleted ESCs form aggressive hemorrhagic teratomas with increased endoderm, reduced neuroectoderm, and ectopic appearance of trophoblastic giant cells. Thus, 5hmC is an epigenetic modification associated with the pluripotent state, and Tet1 functions to regulate the lineage differentiation potential of ESCs.
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http://dx.doi.org/10.1016/j.stem.2011.01.008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3134318PMC
February 2011

Impaired hydroxylation of 5-methylcytosine in myeloid cancers with mutant TET2.

Nature 2010 Dec;468(7325):839-43

Department of Pathology, Harvard Medical School, Immune Disease Institute and Program in Cellular and Molecular Medicine, Children’s Hospital Boston, Boston, Massachusetts 02115, USA.

TET2 is a close relative of TET1, an enzyme that converts 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) in DNA. The gene encoding TET2 resides at chromosome 4q24, in a region showing recurrent microdeletions and copy-neutral loss of heterozygosity (CN-LOH) in patients with diverse myeloid malignancies. Somatic TET2 mutations are frequently observed in myelodysplastic syndromes (MDS), myeloproliferative neoplasms (MPN), MDS/MPN overlap syndromes including chronic myelomonocytic leukaemia (CMML), acute myeloid leukaemias (AML) and secondary AML (sAML). We show here that TET2 mutations associated with myeloid malignancies compromise catalytic activity. Bone marrow samples from patients with TET2 mutations displayed uniformly low levels of 5hmC in genomic DNA compared to bone marrow samples from healthy controls. Moreover, small hairpin RNA (shRNA)-mediated depletion of Tet2 in mouse haematopoietic precursors skewed their differentiation towards monocyte/macrophage lineages in culture. There was no significant difference in DNA methylation between bone marrow samples from patients with high 5hmC versus healthy controls, but samples from patients with low 5hmC showed hypomethylation relative to controls at the majority of differentially methylated CpG sites. Our results demonstrate that Tet2 is important for normal myelopoiesis, and suggest that disruption of TET2 enzymatic activity favours myeloid tumorigenesis. Measurement of 5hmC levels in myeloid malignancies may prove valuable as a diagnostic and prognostic tool, to tailor therapies and assess responses to anticancer drugs.
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http://dx.doi.org/10.1038/nature09586DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3003755PMC
December 2010

The behaviour of 5-hydroxymethylcytosine in bisulfite sequencing.

PLoS One 2010 Jan 26;5(1):e8888. Epub 2010 Jan 26.

Department of Pathology, Harvard Medical School and Immune Disease Institute, Boston, Massachusetts, United States of America.

Background: We recently showed that enzymes of the TET family convert 5-mC to 5-hydroxymethylcytosine (5-hmC) in DNA. 5-hmC is present at high levels in embryonic stem cells and Purkinje neurons. The methylation status of cytosines is typically assessed by reaction with sodium bisulfite followed by PCR amplification. Reaction with sodium bisulfite promotes cytosine deamination, whereas 5-methylcytosine (5-mC) reacts poorly with bisulfite and is resistant to deamination. Since 5-hmC reacts with bisulfite to yield cytosine 5-methylenesulfonate (CMS), we asked how DNA containing 5-hmC behaves in bisulfite sequencing.

Methodology/principal Findings: We used synthetic oligonucleotides with different distributions of cytosine as templates for generation of DNAs containing C, 5-mC and 5-hmC. The resulting DNAs were subjected in parallel to bisulfite treatment, followed by exposure to conditions promoting cytosine deamination. The extent of conversion of 5-hmC to CMS was estimated to be 99.7%. Sequencing of PCR products showed that neither 5-mC nor 5-hmC undergo C-to-T transitions after bisulfite treatment, confirming that these two modified cytosine species are indistinguishable by the bisulfite technique. DNA in which CMS constituted a large fraction of all bases (28/201) was much less efficiently amplified than DNA in which those bases were 5-mC or uracil (the latter produced by cytosine deamination). Using a series of primer extension experiments, we traced the inefficient amplification of CMS-containing DNA to stalling of Taq polymerase at sites of CMS modification, especially when two CMS bases were either adjacent to one another or separated by 1-2 nucleotides.

Conclusions: We have confirmed that the widely used bisulfite sequencing technique does not distinguish between 5-mC and 5-hmC. Moreover, we show that CMS, the product of bisulfite conversion of 5-hmC, tends to stall DNA polymerases during PCR, suggesting that densely hydroxymethylated regions of DNA may be underrepresented in quantitative methylation analyses.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0008888PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2811190PMC
January 2010

Prediction of novel families of enzymes involved in oxidative and other complex modifications of bases in nucleic acids.

Cell Cycle 2009 Jun 27;8(11):1698-710. Epub 2009 Jun 27.

National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, MD 20894, USA.

Modified bases in nucleic acids present a layer of information that directs biological function over and beyond the coding capacity of the conventional bases. While a large number of modified bases have been identified, many of the enzymes generating them still remain to be discovered. Recently, members of the 2-oxoglutarate- and iron(II)-dependent dioxygenase super-family, which modify diverse substrates from small molecules to biopolymers, were predicted and subsequently confirmed to catalyze oxidative modification of bases in nucleic acids. Of these, two distinct families, namely the AlkB and the kinetoplastid base J binding proteins (JBP) catalyze in situ hydroxylation of bases in nucleic acids. Using sensitive computational analysis of sequences, structures and contextual information from genomic structure and protein domain architectures, we report five distinct families of 2-oxoglutarate- and iron(II)-dependent dioxygenase that we predict to be involved in nucleic acid modifications. Among the DNA-modifying families, we show that the dioxygenase domains of the kinetoplastid base J-binding proteins belong to a larger family that includes the Tet proteins, prototyped by the human oncogene Tet1, and proteins from basidiomycete fungi, chlorophyte algae, heterolobosean amoeboflagellates and bacteriophages. We present evidence that some of these proteins are likely to be involved in oxidative modification of the 5-methyl group of cytosine leading to the formation of 5-hydroxymethylcytosine. The Tet/JBP homologs from basidiomycete fungi such as Laccaria and Coprinopsis show large lineage-specific expansions and a tight linkage with genes encoding a novel and distinct family of predicted transposases, and a member of the Maelstrom-like HMG family. We propose that these fungal members are part of a mobile transposon. To the best of our knowledge, this is the first report of a eukaryotic transposable element that encodes its own DNA-modification enzyme with a potential regulatory role. Through a wider analysis of other poorly characterized DNA-modifying enzymes we also show that the phage Mu Mom-like proteins, which catalyze the N6-carbamoylmethylation of adenines, are also linked to diverse families of bacterial transposases, suggesting that DNA modification by transposable elements might have a more general presence than previously appreciated. Among the other families of 2-oxoglutarate- and iron(II)-dependent dioxygenases identified in this study, one which is found in algae, is predicted to mainly comprise of RNA-modifying enzymes and shows a striking diversity in protein domain architectures suggesting the presence of RNA modifications with possibly unique adaptive roles. The results presented here are likely to provide the means for future investigation of unexpected epigenetic modifications, such as hydroxymethyl cytosine, that could profoundly impact our understanding of gene regulation and processes such as DNA demethylation.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2995806PMC
http://dx.doi.org/10.4161/cc.8.11.8580DOI Listing
June 2009

Conversion of 5-methylcytosine to 5-hydroxymethylcytosine in mammalian DNA by MLL partner TET1.

Science 2009 May 16;324(5929):930-5. Epub 2009 Apr 16.

Department of Pathology, Harvard Medical School and Immune Disease Institute, 200 Longwood Avenue, Boston, MA 02115, USA.

DNA cytosine methylation is crucial for retrotransposon silencing and mammalian development. In a computational search for enzymes that could modify 5-methylcytosine (5mC), we identified TET proteins as mammalian homologs of the trypanosome proteins JBP1 and JBP2, which have been proposed to oxidize the 5-methyl group of thymine. We show here that TET1, a fusion partner of the MLL gene in acute myeloid leukemia, is a 2-oxoglutarate (2OG)- and Fe(II)-dependent enzyme that catalyzes conversion of 5mC to 5-hydroxymethylcytosine (hmC) in cultured cells and in vitro. hmC is present in the genome of mouse embryonic stem cells, and hmC levels decrease upon RNA interference-mediated depletion of TET1. Thus, TET proteins have potential roles in epigenetic regulation through modification of 5mC to hmC.
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http://dx.doi.org/10.1126/science.1170116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2715015PMC
May 2009

Mouse Eri1 interacts with the ribosome and catalyzes 5.8S rRNA processing.

Nat Struct Mol Biol 2008 May 27;15(5):523-30. Epub 2008 Apr 27.

Immune Disease Institute, Harvard Medical School, 200 Longwood Avenue, Boston, Massachusetts 02115, USA.

Eri1 is a 3'-to-5' exoribonuclease conserved from fission yeast to humans. Here we show that Eri1 associates with ribosomes and ribosomal RNA (rRNA). Ribosomes from Eri1-deficient mice contain 5.8S rRNA that is aberrantly extended at its 3' end, and Eri1, but not a catalytically inactive mutant, converts this abnormal 5.8S rRNA to the wild-type form in vitro and in cells. In human and murine cells, Eri1 localizes to the cytoplasm and nucleus, with enrichment in the nucleolus, the site of preribosome biogenesis. RNA binding residues in the Eri1 SAP and linker domains promote stable association with rRNA and thereby facilitate 5.8S rRNA 3' end processing. Taken together, our findings indicate that Eri1 catalyzes the final trimming step in 5.8S rRNA processing, functionally and spatially connecting this regulator of RNAi with the basal translation machinery.
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http://dx.doi.org/10.1038/nsmb.1417DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3032500PMC
May 2008

The histone H3K4 demethylase SMCX links REST target genes to X-linked mental retardation.

Nature 2007 May 29;447(7144):601-5. Epub 2007 Apr 29.

Division of Endocrinology, Diabetes, and Hypertension, Department of Medicine and BCMP, Brigham and Women's Hospital and Harvard Medical School, 221 Longwood Avenue Boston, Massachusetts 02115, USA.

Gene transcription is critically influenced by chromatin structure and the modification status of histone tails. Methylation of lysine residues in histone tails is dynamically regulated by the opposing activities of histone methyltransferases and histone demethylases. Here we show that JARID1C/SMCX, a JmjC-domain-containing protein implicated in X-linked mental retardation and epilepsy, possesses H3K4 tri-demethylase activity and functions as a transcriptional repressor. An SMCX complex isolated from HeLa cells contains additional chromatin modifiers (the histone deacetylases HDAC1 and HDAC2, and the histone H3K9 methyltransferase G9a) and the transcriptional repressor REST, suggesting a direct role for SMCX in chromatin dynamics and REST-mediated repression. Chromatin immunoprecipitation reveals that SMCX and REST co-occupy the neuron-restrictive silencing elements in the promoters of a subset of REST target genes. RNA-interference-mediated depletion of SMCX derepresses several of these targets and simultaneously increases H3K4 trimethylation at the sodium channel type 2A (SCN2A) and synapsin I (SYN1) promoters. We propose that loss of SMCX activity impairs REST-mediated neuronal gene regulation, thereby contributing to SMCX-associated X-linked mental retardation.
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http://dx.doi.org/10.1038/nature05823DOI Listing
May 2007

PtdIns(4,5)P2 functions at the cleavage furrow during cytokinesis.

Curr Biol 2005 Aug;15(15):1407-12

Division of Signal Transduction, Beth Israel-Deaconess Medical Center, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA.

Phosphoinositides play important roles in regulating the cytoskeleton and vesicle trafficking, potentially important processes at the cleavage furrow. However, it remains unclear which, if any, of the phosphoinositides play a role during cytokinesis. A systematic analysis to determine if any of the phosphoinositides might be present or of functional importance at the cleavage furrow has not been published. Several studies hint at a possible role for one or more phosphoinositides at the cleavage furrow. The best of these are genetic data identifying mutations in phosphoinositide-modifying enzymes (a PtdIns(4)P-5-kinase in S. pombe and a PI-4-kinase in D. melanogaster) that interfere with cytokinesis. The genetic nature of these experiments leaves questions as to how direct may be their contribution to cytokinesis. Here we show that a single phosphoinositide, PtdIns(4,5)P2, specifically accumulates at the furrow. Interference with PtdIns(4,5)P2 interferes with adhesion of the plasma membrane to the contractile ring at the furrow. Finally, four distinct interventions to specifically interfere with PtdIns(4,5)P2 each impair cytokinesis. We conclude that PtdIns(4,5)P2 is present at the cleavage furrow and is required for normal cytokinesis at least in part because of a role in adhesion between the contractile ring and the plasma membrane.
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http://dx.doi.org/10.1016/j.cub.2005.06.059DOI Listing
August 2005