Genome-wide alteration of 5-hydroxymenthylcytosine in a mouse model of Alzheimer’s disease
- Liqi Shu†1,
- Wenjia Sun†2,
- Liping Li†3, 4,
- Zihui Xu1, 5,
- Li Lin1,
- Pei Xie3, 4,
- Hui Shen3, 4,
- Luoxiu Huang1,
- Qi Xu6,
- Peng Jin1Email author and
- Xuekun Li3, 4Email author
© Shu et al. 2016
Received: 22 October 2015
Accepted: 12 May 2016
Published: 20 May 2016
Alzheimer’s disease (AD) is the most common form of neurodegenerative disorder that leads to a decline in cognitive function. In AD, aggregates of amyloid β peptide precede the accumulation of neurofibrillary tangles, both of which are hallmarks of the disease. The great majority (>90 %) of the AD cases are not originated from genetic defects, therefore supporting the central roles of epigenetic modifications that are acquired progressively during the life span. Strong evidences have indicated the implication of epigenetic modifications, including histone modification and DNA methylation, in AD. Recent studies revealed that 5-hydroxymethylcytosine (5hmC) is dynamically regulated during neurodevelopment and aging.
We show that amyloid peptide 1–42 (Aβ1-42) could significantly reduce the overall level of 5hmC in vitro. We found that the level of 5hmC displayed differential response to the pathogenesis in different brain regions, including the cortex, cerebellum, and hippocampus of APP-PSEN1 double transgenic (DTg) mice. We observed a significant decrease of overall 5hmC in hippocampus, but not in cortex and cerebellum, as the DTg mice aged. Genome-wide profiling identified differential hydroxymethylation regions (DhMRs) in DTg mice, which are highly enriched in introns, exons and intergenic regions. Gene ontology analyses indicated that DhMR-associated genes are highly enriched in multiple signaling pathways involving neuronal development/differentiation and neuronal function/survival.
5hmC-mediated epigenetic regulation could potentially be involved in the pathogenesis of AD.
Alzheimer’s disease (AD) is a progressive neurodegenerative disease involving multiple pathologic processes and is characterized by the deposition of amyloid beta (Aβ) peptide, neurofibrillary tangles (NFTs) composed of hyperphosphorylated protein tau, and neuronal cell death [1, 2]. Recent studies indicate that epigenetic pathways could be involved in the pathogenesis of AD [3, 4]. DNA methylation (5-methylcytosine, 5mC) plays important roles in regulating gene expression and is involved in multiple neurodevelopmental and neurodegenerative disorders [5–7].
Changes in 5mC at the global level or at specific loci are seen in the brain tissues of AD model mice, as well as AD patients [4, 8–12]. Although some regions and loci show hypermethylation , global DNA hypomethylation has been observed in the entorhinal cortex of some AD patients , suggesting DNA methylation is differentially affected in a region- and loci-specific manner. Previous studies also found that the promoter regions of amyloid precursor protein (APP) and presenilin 1 (PSEN1) displayed age-dependent hypomethylation [10, 15–17]. Furthermore, in vitro hypomethylation of PSEN1 increased the cleavage of APP and the production of Aβ in a neuroblastoma cell line . Recently, two large-scale epigenome-wide association studies uncovered the alteration of site-specific methylation in the brains of AD patients [11, 12]. These results imply DNA methylation could play important roles in the pathogenesis of AD.
Recently, another cytosine modification, 5-hydroxymethylcytosine (5hmC), was identified and found to be highly abundant in the neuronal system [19–21]. Ten-eleven translocation (Tet) family proteins, including Tet1, Tet2, and Tet3, are known to catalyze the hydroxylation of 5mC to 5hmC [19, 22–24]. Recent studies strongly indicate that 5hmC not only serves as an intermediate of DNA demethylation, but can also perform as a stable epigenetic marker. 5hmC is ~10-fold more enriched in neurons than other cell types, and it is acquired globally and exhibits dynamic features and region-specific patterns during postnatal development and aging of the neuronal system [20, 25, 26]. Genome-wide studies reveal that 5hmC can be enriched in distinct genomic regions, such as gene bodies, promoters, and distal regulatory regions [27–29], and the enrichment of 5hmC ate gene bodies could be positively correlated with transcriptional level, which might be achieved via interaction with histone modifications [30–34]. The alteration of global 5hmC and differentially hydroxymethylated regions (DhMRs) are seen in several neurodevelopmental diseases, including Rett syndrome, autism, and neurodegenerative diseases like Huntington’s disease and fragile X-associated tremor/ataxia syndrome (FXTAS), suggesting 5hmC could play important roles in neurological diseases [20, 25, 35–37].
Despite the clear alteration of DNA methylation observed in AD, whether and how 5hmC is involved in AD pathogenesis still remain largely unknown. Using an immunostaining method, Condliffe et al. found a significant decrease of global 5hmC in the cortex and cerebellum of AD patients . In contrast, using the same technique, other studies reported an increase of global 5hmC in both AD mouse model and patients samples [39–41]. To study the alteration of 5hmC in AD and explore the potential role(s) of 5hmC-mediated epigenetic regulation in the pathogenesis of AD, here we investigated the effect of Aβ on 5hmC in vitro and found Aβ treatment could significantly decrease the level of 5hmC in a dose-dependent pattern. Furthermore, we found that 5hmC levels displayed an age-dependent decrease in the hippocampus, but not in the cortex and cerebellum, of APP-PSEN1 double-transgenic (DTg) mice. Using a chemical-labeling 5hmC enrichment approach, we performed genome-wide profiling of 5hmC. We found that, although AD pathogenesis did not change the overall distribution of 5hmC, there were differentially hydroxymethylated regions (DhMRs) in DTg mice. The DhMRs identified are involved in a number of neuronal signaling pathways, indicating a 5hmC-mediated epigenetic pathway could potentially play important roles in the pathogenesis of AD.
Aβ reduces the global level of 5-hydroxymethylcytosine in vitro
Considering cognitive function is severely impaired and the roles of adult neurogenesis in learning and memory, we further tested the effect of Aβ peptide on 5hmC levels in adult neural stem cells (aNSCs). ANSCs harbors in specific regions, subventricular zone of lateral ventricle and subgranular zone of dentate gyrus of adult mammalian brain, and is involved in neurological disorders including AD. The isolated aNSCs were positive for neural stem cell markers Nestin and Sox2 (Fig. 1c-f). After treated with Aβ peptide at a 1-μM concentration for 48 h, the level of 5hmC was also significantly decreased in the cultured aNSCs (Fig. 1g-h). We also compared the overall level of 5hmC in HEK293ft cells, aNSCs and neuronal tissues. We found that 5hmC level was significantly higher in neuronal tissues than in HEK293ft and aNSCs cells, which both are capable of proliferation (Fig. i-j). Taken together, these results indicate that Aβ peptide could significantly affect the level of 5hmC in multiple cultured cells.
5-hydroxymethylcytosine level decreases during aging in an AD mouse model
Acquisition of 5hmC on gene bodies is altered in aged AD mice
We next determined the distribution of 5hmC on distinct genomic regions. Genome-wide 5hmC read density was detected in the DTg mouse model (Fig. 3c, d). At the 12-week point, we saw no significant difference between WT and DTg mice (Fig. 3c). However, until the 67-week point, the shifted entire plot pattern suggested that WT bins had more 5hmC reads (Fig. 3d). Furthermore, the distribution of 5hmC was studied at 2.5 kb up- and downstream of transcription end sites (TESs), transcription starting sites (TSSs), gene bodies, and CpG islands by ngs-plot software (Fig. 3e-h). We found that the distribution of 5hmC showed no observable differences on TESs and CpG islands between WT and DTg mice (Fig. 3e, h), whereas it was slightly increased on TSSs in WT mice compared to age-matched DTg mice (Fig. 3f). From the 12-week to 67-week time point, 5hmC was acquired on gene bodies in both WT and DTg mice (Fig. 3g), however, at 12- and 67-week time points, the enrichment of 5hmC in gene bodies showed a significant decrease in DTg mice compared to age-matched WT mice (t-test, p < 0.0001), suggesting the acquisition of 5hmC in gene bodies was inhibited during AD pathogenesis.
Differential hydroxymethylated regions (DhMRs) associated with AD
To further reveal the biological function of identified DhMRs in both adult and aged DTg mice, the genes associated with these DhMRs were extracted for enrichment analysis. We found 167 genes associating with the identified DTg-specific DhMRs (Additional file 3: Table S2). DhMR-associated genes were highly enriched in multiple signaling pathways, such as the Wnt and ErbB pathways, which play important roles in the neuronal system (Fig. 4c). Furthermore, we generated 4557 aged DTg mice-specific peaks, and extracted 2424 genes, which associated with those peaks (Additional file 4: Table S3). Gene Ontology assay showed that those genes significantly enriched in some pathways, such as Alzheimer’s disease pathway, Wnt signalling, etc. (Fig. 4d). Taken together, these results suggest that the specific enrichment of 5hmC could play some role(s) in regulating the expression of genes related neuronal function, and involve in the pathogenesis of Alzheimer’s disease.
Previous studies also noted that the enrichment of 5hmC in gene bodies might be positively correlated with gene expression . In our AD model mice, two AD-associated genes, APP and PSEN1, were over-expressed. We found 5hmC peaks highly enriched in APP gene bodies, especially in exons, in both adult and aged DTg mice compared to WT mice (Fig. 5d, over two-fold difference, p < 1x10−5). It is of interest to note that we did not see significant difference of 5hmC distribution in PSEN1. These data indicated that 5hmC enrichment could be one of the mechanisms promoting gene expression.
In the present study, we performed in vitro and in vivo studies to characterize the alterations of 5hmC-mediated DNA demethylation in a mouse model of AD. We found the overall level of 5hmC is significantly higher in brain tissues than in cell lines and adult neural stem cells. AD pathogenic protein amyloid peptide led to a decrease of global 5hmC in cultured cells. Our in vivo study also found the level of 5hmC decreased in one specific brain region, i.e., the hippocampus, but not other studied brain regions of AD mice during the pathogenesis of AD. Genome-wide profiling results indicated that the distribution of 5hmC altered in distinct genomic regions, especially in gene bodies. The differentially hydroxymethylated regions (DhMRs) identified in hippocampus of aged D-Tg mice displayed high enrichment of multiple signaling pathways that are related to neuronal development and neuronal function. Some AD-associated genes showed altered hydroxymethylation. Our study therefore uncovered new roles for 5hmC-mediated epigenetic modification in neurologic disorders and revealed a new layer of the pathogenic mechanism of AD.
Global and site-specific alterations of DNA methylation had been identified in AD [10, 14, 15, 43–45]. Epigenome-wide association with AD revealed the relationship between differential methylation of CpGs and the expression of nearby genes, some of which are connected to a known AD susceptibility network [11, 12]. Previous studies had inconsistently reported about the alteration of DNA demethylation [38–41]. In our present study, Aβ treatment led to a significant decrease in global 5hmC level of different types of cells in vitro. The significant alteration of demethyaltion was only observed in hippocampus but not in cortex and cerebellum during the ageing of AD model mice, suggesting the alteration of demethylation is region specific, which might explain the data contradiction between different studies. Considering the acquired Aβ deposition during the ageing of AD model mice, these results indicated the increased Aβ deposition decreased overall 5hmC level. Together with previous studies, it supports the idea that dysregulation of DNA demethylation is age- and region/loci specific, indicating an interaction between amyloid accumulation and DNA modification.
Previous studies have found 5hmC is highly enriched in the neuronal system [19, 26], and the enrichment is enhanced and displays dynamic features during postnatal development and aging of the brain [20, 46], indicating 5hmC could be important for brain function. Subsequent studies did find that 5hmC-mediated epigenetic modification is involved in multiple neurological disorders, including autism spectrum disorders, Huntington’s disease, and FXTAS [20, 25, 36, 37]. Moreover, the Rett syndrome protein MeCP2 could bind to 5hmC, and its dosage is negatively correlated with 5hmC level [20, 35]. Our present results revealed that the overall distribution features of 5hmC in TSSs, TESs, and CpG islands did not change, but the enrichment of 5hmC in gene bodies was significantly decreased compared with age-matched WT mice. Previous research found that the 5hmC level in gene bodies is positively correlated with gene expression [20, 47]. Our results that the highly enrichment of 5hmC in the gene body of APP further supports this concept. Considering no observable difference of 5hmC in another overexpressed gene PSEN1, it suggests that 5hmC enrichment in gene body is one but all of mechanisms to modulate gene expression. Further, our present studies identified some DhMR-associated genes enriched in multiple signaling pathways that are related to neuronal function and neurological disorders [20, 25, 36, 37]. These results suggest a potential mechanism to explain how 5hmC-mediated epigenetic modification functions.
Our present studies also uncovered altered demethylation in some specific loci in aged D-Tg mice, and some loci were related with the onset and progress of AD. Interestingly, the loci with altered DNA demethylation identified in our study were also found displaying altered DNA methylation profile by two large-scale studies in human [11, 12]. Although our present and these two studies mainly provided the relevant evidence, all these results suggest DNA methylation and demethylation changes are involved in AD. Further experimental work needs to be conducted to address the mechanism how altered DNA methylation and demethylation affect the onset and pathogenesis of AD.
Overall, our results indicate that not only is the global level but also the distribution features of 5hmC altered in AD model mice. Aβ treatment decreased 5hmC both in vitro and in vivo. The acquisition of 5hmC in gene bodies during postnatal development was significantly inhibited in the hippocampus of AD model mice, although the overall features in TSSs, TESs, and CpG islands were unaffected. The DhMR-associated genes identified in AD mice are specifically related to some signaling pathways that play a role in neuronal function and neurological disorders. Taken together, our present results argue that 5hmC-mediated epigenetic modification could have an important function in AD.
Twelve- and 67-week-old wild-type (WT) and APP/PS1 double transgenic littermate mice were used in this study . Mice were maintained at ambient temperature (22-24 °C) on a 12:12 light/dark cycle with free access to food and water. All animal procedures were performed according to protocols approved by Emory University Institutional Animal Care and Use Committee.
Genomic DNA isolation and 5hmC dot-blot
Genomic DNA was extracted as described previously . Briefly, the dissected brain samples or cells were homogenized in lysis buffer (5 mM EDTA, 0.2 % SDS, 200 mM NaCl in 100 mM Tris–HCl, pH 8.5) supplemented with proteinase K, and samples were kept at 56 °C overnight. The second day, an equal volume of phenol:chloroform:isoamyl alcohol (25:24:1, P-3803, Sigma) was added, mixed completely, and centrifuged at 14,000 rpm for 10 min. An equal volume of isopropanol was added to the supernatant to precipitate DNA, which was dissolved with 10 mM Tris–HCl (pH 8.0).
5hmC dot-blot was performed as before . In brief, genomic DNA was spotted on an Amersham Hybond-N+ membrane (GE Healthcare), followed by baking at 80 °C for 30 min. The membrane was incubated with polyclonal 5hmC antibody (Active Motif, #39769) overnight at 4 °C. The second day, a horseradish-peroxidase-conjugated secondary antibody against rabbit was used to probe.
Cell culture and Aβ treatment
HEK293 cells were maintained in DMEM supplemented with 10 % fetal bovine serum, 2 mM glutamine, and 100 U penicillin–streptomycin at 37 °C in a humidified incubator containing 5 % CO2. Cells were treated with Aβ peptide (Sigma, A9810) at a concentration of 0.5, 0.75, or 1 μM for 48 h, respectively. The isolation, culture and determination of adult neural stem cells were performed as described previously .
The cultured aNSCs were fixed with 4 % parafromaldehyde for 30 min at room temperature, followed by washing with cold PBS for 15 min. The cells were blocked with 3 % goat serum and 0.1 % TritonX-100 in PBS for 1 h at room temperature, followed by the incubation with primary antibodies at 4 °C overnight. The second day, the cells were incubated with secondary antibodies after washed with PBS for 30 min. The images were taken with a Zeiss confocal microscope. Primary antibodies: Rabbit SOX2 (Millipore, Ab5603), Mouse Nestin (BD, #556309). Secondary antibodies: goat anti Rabbit 488 (Invitrogen, A11008), goat anti Mouse 568 (Invitrogen, A11004).
5hmC-specific enrichment and high-throughput sequencing
Chemical labeling-based 5hmC enrichment was described previously . Briefly, DNA was sonicated to 100–500 bp, and then mixed with 100 μl solution containing 50 mM HEPES buffer (pH 7.9), 25 mM MgCl2, 250 μM UDP-6-N3-Glu, and 2.25 μM β-glucosyltransferase for 1 h at 37 °C. DNA substrates were purified via Qiagen DNA purification kit. 150 μM dibenzocyclooctyne modified biotin was then added to the purified DNA, and the labeling reaction was performed for 2 h at 37 °C. The biotin-labeled DNA was enriched by Streptavidin-coupled Dynabeads (Dynabeads® MyOne™ Streptavidin T1, Life Technologies) and purified.
5hmC libraries were generated with 25 ng input or 5hmC-captured DNA according to the manufacturer’s protocol (NEBNext ChIP-Seq Library Prep Reagent Set for Illumina). DNA fragments between 150 and 300 bp were gel-purified after the adapter ligation step. An Agilent 2100 BioAnalyzer was used to quantify the amplified DNA. 20 pM diluted libraries were eventually used for sequencing.
Sequence alignment and mapped reads annotation
FASTQ sequence files were aligned to mouse NCBI37v1/mm9 references using Bowtie 0.12.9. Each unique mapped read with no more than two mismatches in the first 25 bp was concatenated to achieve combined wild-type and APP/PS1 mice 5hmC sequence. Association of mapped reads with genomic features was performed by overlapping reads files with known genomic features obtained from UCSC Tables for NCBI37v1/mm9. Unique 5hmC mapped reads were plotted to various genomic regions using an R program package termed ngsplot (https://code.google.com/p/ngsplot/).
DhMR identification, annotation, and motif analysis
Model-based Analysis of ChIP-Seq (MACS) software  was adopted to identify DhMRs between WT and DTg mice by directly comparing one to the other, rather than comparing to the input. The effective genome size = 1.87 × 109, tag size = 38, bandwidth = 200, P-value cutoff = 1.00 × 10−5. Identified WT and DTg-specific DhMRs were annotated to various genomic regions and associated genes by HOMER software . DhMR-associated genes were extracted, and enrichment analysis was performed with WebGestalt (http://bioinfo.vanderbilt.edu/webgestalt/) .
Data are expressed as the mean ± standard error of the mean (s. e. m.), and statistical significance of differences between different groups was assessed using the t-test or ANOVA assay.
We thank Ms. C. Strauss for critical reading of the manuscript. X.L. was supported by the National Natural Science Foundation of China (31371309) and National Key Basic Research Program of China (No.2014CB943001). P.J. was supported in part by National Institutes of Health (NS079625 and P50AG025688 to PJ). Z.X. was supported by the National Natural Science Foundation of China (31400916).
XL and PJ conceived and designed the study. LS, WS, LL, ZX, PX, LL, HS, LH and QX performed the experiments. XL, PJ wrote the manuscript. All authors read and approved the final manuscript.
The authors declare they have no competing interests.
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- Duyckaerts C, Delatour B, Potier MC. Classification and basic pathology of Alzheimer disease. Acta Neuropathol. 2009;118:5–36.View ArticlePubMedGoogle Scholar
- Finder VH. Alzheimer’s disease: a general introduction and pathomechanism. J Alzheimers Dis. 2010;22 Suppl 3:5–19.PubMedGoogle Scholar
- Gapp K, Woldemichael BT, Bohacek J, Mansuy IM. Epigenetic regulation in neurodevelopment and neurodegenerative diseases. Neuroscience. 2014;264:99–111.View ArticlePubMedGoogle Scholar
- Chouliaras L, Rutten BP, Kenis G, Peerbooms O, Visser PJ, Verhey F, van Os J, Steinbusch HW, van den Hove DL. Epigenetic regulation in the pathophysiology of Alzheimer’s disease. Prog Neurobiol. 2010;90:498–510.View ArticlePubMedGoogle Scholar
- Bergman Y, Cedar H. DNA methylation dynamics in health and disease. Nat Struct Mol Biol. 2013;20:274–81.View ArticlePubMedGoogle Scholar
- Jakovcevski M, Akbarian S. Epigenetic mechanisms in neurological disease. Nat Med. 2012;18:1194–204.View ArticlePubMedPubMed CentralGoogle Scholar
- Jaenisch R, Bird A. Epigenetic regulation of gene expression: how the genome integrates intrinsic and environmental signals. Nat Genet. 2003;33(Suppl):245–54.View ArticlePubMedGoogle Scholar
- Chouliaras L, Mastroeni D, Delvaux E, Grover A, Kenis G, Hof PR, Steinbusch, HW, Coleman PD, Rutten BP, van den Hove DL. Consistent decrease in global DNA methylation and hydroxymethylation in the hippocampus of Alzheimer’s disease patients. Neurobiol Aging. 2013;34:2091–9.View ArticlePubMedPubMed CentralGoogle Scholar
- Coppieters N, Dragunow M. Epigenetics in Alzheimer’s disease: a focus on DNA modifications. Curr Pharm Des. 2011;17:3398–412.View ArticlePubMedGoogle Scholar
- Rao JS, Keleshian VL, Klein S, Rapoport SI. Epigenetic modifications in frontal cortex from Alzheimer’s disease and bipolar disorder patients. Transl Psychiatry. 2012;2:e132.View ArticlePubMedPubMed CentralGoogle Scholar
- De Jager PL, Srivastava G, Lunnon K, Burgess J, Schalkwyk LC, Yu L, Eaton ML, Keenan BT, Ernst J, McCabe C, et al. Alzheimer’s disease: early alterations in brain DNA methylation at ANK1, BIN1, RHBDF2 and other loci. Nat Neurosci. 2014;17:1156–63.View ArticlePubMedPubMed CentralGoogle Scholar
- Lunnon K, Smith R, Hannon E, De Jager PL, Srivastava G, Volta M, Troakes C, Al-Sarraj S, Burrage J, Macdonald R, et al. Methylomic profiling implicates cortical deregulation of ANK1 in Alzheimer’s disease. Nat Neurosci. 2014;17:1164–70.View ArticlePubMedPubMed CentralGoogle Scholar
- Siegmund KD, Connor CM, Campan M, Long TI, Weisenberger DJ, Biniszkiewicz D, Jaenisch R, Laird PW, Akbarian S. DNA methylation in the human cerebral cortex is dynamically regulated throughout the life span and involves differentiated neurons. PLoS One. 2007;2:e895.View ArticlePubMedPubMed CentralGoogle Scholar
- Mastroeni D, Grover A, Delvaux E, Whiteside C, Coleman PD, Rogers J. Epigenetic changes in Alzheimer’s disease: decrements in DNA methylation. Neurobiol Aging. 2010;31:2025–37.View ArticlePubMedPubMed CentralGoogle Scholar
- Bakulski KM, Dolinoy DC, Sartor MA, Paulson HL, Konen JR, Lieberman AP, Albin RL, Hu H, Rozek LS. Genome-wide DNA methylation differences between late-onset Alzheimer’s disease and cognitively normal controls in human frontal cortex. J Alzheimers Dis. 2012;29:571–88.PubMedGoogle Scholar
- Barrachina M, Ferrer I. DNA methylation of Alzheimer disease and tauopathy-related genes in postmortem brain. J Neuropathol Exp Neurol. 2009;68:880–91.View ArticlePubMedGoogle Scholar
- Wang SC, Oelze B, Schumacher A. Age-specific epigenetic drift in late-onset Alzheimer’s disease. PLoS One. 2008;3:e2698.View ArticlePubMedPubMed CentralGoogle Scholar
- Fuso A, Seminara L, Cavallaro RA, D’Anselmi F, Scarpa S. S-adenosylmethionine/homocysteine cycle alterations modify DNA methylation status with consequent deregulation of PS1 and BACE and beta-amyloid production. Mol Cell Neurosci. 2005;28:195–204.View ArticlePubMedGoogle Scholar
- Kriaucionis S, Heintz N. The nuclear DNA base 5-hydroxymethylcytosine is present in Purkinje neurons and the brain. Science. 2009;324:929–30.View ArticlePubMedPubMed CentralGoogle Scholar
- Szulwach KE, Li X, Li Y, Song CX, Wu H, Dai Q, Irier H, Upadhyay AK, Gearing M, Levey AI, et al. 5-hmC-mediated epigenetic dynamics during postnatal neurodevelopment and aging. Nat Neurosci. 2011;14:1607–16.View ArticlePubMedPubMed CentralGoogle Scholar
- Jin SG, Wu X, Li AX, Pfeifer GP. Genomic mapping of 5-hydroxymethylcytosine in the human brain. Nucleic Acids Res. 2011;39:5015–24.View ArticlePubMedPubMed CentralGoogle Scholar
- Gu TP, Guo F, Yang H, Wu HP, Xu GF, Liu W, Xie ZG, Shi L, He X, Jin SG, et al. The role of Tet3 DNA dioxygenase in epigenetic reprogramming by oocytes. Nature. 2011;477:606–10.View ArticlePubMedGoogle Scholar
- Ito S, Shen L, Dai Q, Wu SC, Collins LB, Swenberg JA, He C, Zhang Y. Tet proteins can convert 5-methylcytosine to 5-formylcytosine and 5-carboxylcytosine. Science. 2011;333:1300–3.View ArticlePubMedPubMed CentralGoogle Scholar
- Tahiliani M, Koh KP, Shen Y, Pastor WA, Bandukwala H, Brudno Y, Agarwal S, Iyer LM, Liu DR, Aravind L, et al. Conversion of 5-methylcytosine to 5-hydroxymethylcytosine in mammalian DNA by MLL partner TET1. Science. 2009;324:930–5.View ArticlePubMedPubMed CentralGoogle Scholar
- Wang T, Pan Q, Lin L, Szulwach KE, Song CX, He C, Wu H, Warren ST, Jin P, Duan R, et al. Genome-wide DNA hydroxymethylation changes are associated with neurodevelopmental genes in the developing human cerebellum. Hum Mol Genet. 2012;21:5500–10.View ArticlePubMedPubMed CentralGoogle Scholar
- Song CX, Szulwach KE, Fu Y, Dai Q, Yi C, Li X, Li Y, Chen CH, Zhang W, Jian X, et al. Selective chemical labeling reveals the genome-wide distribution of 5-hydroxymethylcytosine. Nat Biotechnol. 2011;29:68–72.View ArticlePubMedPubMed CentralGoogle Scholar
- Ficz G, Branco MR, Seisenberger S, Santos F, Krueger F, Hore TA, Marques CJ, Andrews S, Reik W. Dynamic regulation of 5-hydroxymethylcytosine in mouse ES cells and during differentiation. Nature. 2011;473:398–402.View ArticlePubMedGoogle Scholar
- Pastor WA, Pape UJ, Huang Y, Henderson HR, Lister R, Ko M, McLoughlin EM, Brudno Y, Mahapatra S, Kapranov P, et al. Genome-wide mapping of 5-hydroxymethylcytosine in embryonic stem cells. Nature. 2011;473:394–7.View ArticlePubMedPubMed CentralGoogle Scholar
- Wu H, Zhang Y. Mechanisms and functions of Tet protein-mediated 5-methylcytosine oxidation. Genes Dev. 2011;25:2436–52.View ArticlePubMedPubMed CentralGoogle Scholar
- Szulwach KE, Li X, Li Y, Song CX, Han JW, Kim S, Namburi S, Hermetz K, Kim JJ, Rudd MK, et al. Integrating 5-hydroxymethylcytosine into the epigenomic landscape of human embryonic stem cells. PLoS Genet. 2011;7:e1002154.View ArticlePubMedPubMed CentralGoogle Scholar
- Etchegaray JP, Chavez L, Huang Y, Ross KN, Choi J, Martinez-Pastor B, Walsh RM, Sommer CA, Lienhard M, Gladden A, et al. The histone deacetylase SIRT6 controls embryonic stem cell fate via TET-mediated production of 5-hydroxymethylcytosine. Nat Cell Biol. 2015;17:545–57.View ArticlePubMedPubMed CentralGoogle Scholar
- Choi I, Kim R, Lim HW, Kaestner KH, Won KJ. 5-hydroxymethylcytosine represses the activity of enhancers in embryonic stem cells: a new epigenetic signature for gene regulation. BMC Genomics. 2014;15:670.View ArticlePubMedPubMed CentralGoogle Scholar
- Hahn MA, Qiu R, Wu X, Li AX, Zhang H, Wang J, Jui J, Jin SG, Jiang Y, Pfeifer GP, et al. Dynamics of 5-hydroxymethylcytosine and chromatin marks in Mammalian neurogenesis. Cell Rep. 2013;3:291–300.View ArticlePubMedPubMed CentralGoogle Scholar
- Deplus R, Delatte B, Schwinn MK, Defrance M, Mendez J, Murphy N, Dawson MA, Volkmar M, Putmans P, Calonne E, et al. TET2 and TET3 regulate GlcNAcylation and H3K4 methylation through OGT and SET1/COMPASS. EMBO J. 2013;32:645–55.View ArticlePubMedPubMed CentralGoogle Scholar
- Mellen M, Ayata P, Dewell S, Kriaucionis S, Heintz N. MeCP2 binds to 5hmC enriched within active genes and accessible chromatin in the nervous system. Cell. 2012;151:1417–30.View ArticlePubMedPubMed CentralGoogle Scholar
- Wang F, Yang Y, Lin X, Wang JQ, Wu YS, Xie W, Wang D, Zhu S, Liao YQ, Sun Q, et al. Genome-wide loss of 5-hmC is a novel epigenetic feature of Huntington’s disease. Hum Mol Genet. 2013;22:3641–53.View ArticlePubMedGoogle Scholar
- Yao B, Lin L, Street RC, Zalewski ZA, Galloway JN, Wu H, Nelson DL, Jin P. Genome-wide alteration of 5-hydroxymethylcytosine in a mouse model of fragile X-associated tremor/ataxia syndrome. Hum Mol Genet. 2013; in press.Google Scholar
- Condliffe D, Wong A, Troakes C, Proitsi P, Patel Y, Chouliaras L, Fernandes C, Cooper J, Lovestone S, Schalkwyk L, et al. Cross-region reduction in 5-hydroxymethylcytosine in Alzheimer’s disease brain. Neurobiol Aging. 2014;35:1850–4.View ArticlePubMedPubMed CentralGoogle Scholar
- Bradley-Whitman MA, Lovell MA. Epigenetic changes in the progression of Alzheimer’s disease. Mech Ageing Dev. 2013;134:486–95.View ArticlePubMedGoogle Scholar
- Coppieters N, Dieriks BV, Lill C, Faull RL, Curtis MA, Dragunow M. Global changes in DNA methylation and hydroxymethylation in Alzheimer’s disease human brain. Neurobiol Aging. 2014;35:1334–44.View ArticlePubMedGoogle Scholar
- Cadena-Del-Castillo C, Valdes-Quezada C, Carmona-Aldana F, Arias C, Bermudez-Rattoni F, Recillas-Targa F. Age-dependent increment of hydroxymethylation in the brain cortex in the triple-transgenic mouse model of Alzheimer’s disease. J Alzheimers Dis. 2014; in press.Google Scholar
- Zhang Y, Liu T, Meyer CA, Eeckhoute J, Johnson DS, Bernstein BE, Nusbaum C, Myers RM, Brown M, Li W, et al. Model-based analysis of ChIP-Seq (MACS). Genome Biol. 2008;9:R137.View ArticlePubMedPubMed CentralGoogle Scholar
- Mastroeni D, McKee A, Grover A, Rogers J, Coleman PD. Epigenetic differences in cortical neurons from a pair of monozygotic twins discordant for Alzheimer’s disease. PLoS One. 2009;4:e6617.View ArticlePubMedPubMed CentralGoogle Scholar
- Van den Hove DL, Kompotis K, Lardenoije R, Kenis G, Mill J, Steinbusch HW, Lesch KP, Fitzsimons CP, De Strooper B, Rutten BP. Epigenetically regulated microRNAs in Alzheimer’s disease. Neurobiol Aging. 2014;35:731–45.View ArticlePubMedGoogle Scholar
- Bennett DA, Yu L, Yang J, Srivastava GP, Aubin C, De Jager PL. Epigenomics of Alzheimer’s disease. Transl Res. 2014; in press.Google Scholar
- Lister R, Mukamel EA, Nery JR, Urich M, Puddifoot CA, Johnson ND, Lucero J, Huang Y, Dwork AJ, Schultz MD, et al. Global epigenomic reconfiguration during mammalian brain development. Science. 2013;341:1237905.View ArticlePubMedPubMed CentralGoogle Scholar
- Colquitt BM, Allen WE, Barnea G, Lomvardas S. Alteration of genic 5-hydroxymethylcytosine patterning in olfactory neurons correlates with changes in gene expression and cell identity. Proc Natl Acad Sci U S A. 2013;110:14682–7.View ArticlePubMedPubMed CentralGoogle Scholar
- Kummer MP, Hammerschmidt T, Martinez A, Terwel D, Eichele G, Witten A, Figura S, Stoll M, Schwartz S, Pape HC, et al. Ear2 deletion causes early memory and learning deficits in APP/PS1 mice. J Neurosci. 2014;34:8845–54.View ArticlePubMedPubMed CentralGoogle Scholar
- Li X, Barkho BZ, Luo Y, Smrt RD, Santistevan NJ, Liu C, Kuwabara T, Gage FH, Zhao X. Epigenetic regulation of the stem cell mitogen Fgf-2 by Mbd1 in adult neural stem/progenitor cells. J Biol Chem. 2008;283:27644–52.View ArticlePubMedPubMed CentralGoogle Scholar
- Heinz S, Benner C, Spann N, Bertolino E, Lin YC, Laslo P, Cheng JX, Murre C, Singh H, Glass CK. Simple combinations of lineage-determining transcription factors prime cis-regulatory elements required for macrophage and B cell identities. Mol Cell. 2010;38:576–89.View ArticlePubMedPubMed CentralGoogle Scholar
- Zhang B, Kirov S, Snoddy J. WebGestalt: an integrated system for exploring gene sets in various biological contexts. Nucleic Acids Res. 2005;33:W741–8.View ArticlePubMedPubMed CentralGoogle Scholar