Active and Passive Demethylation of Male and Female Pronuclear DNA in the Mammalian Zygote

Guo, Fan; Li, Xianlong; Liang, Dan; Li, Tong; Zhu, Ping; Guo, Hongshan; Wu, Xinglong; Wen, Lu; Gu, Tianpeng; Hu, Boqiang; Walsh, Colum P.; Li, Jinsong; Tang, Fuchou; Xu, Guoliang

doi:10.1016/j.stem.2014.08.003

Cited by 329 publications

(345 citation statements)

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“…The genome-wide DNA demethylation observed in early embryos appears to be a default pathway independent of TET function or oxi-mC modification: both the 5hmC-modified paternal genome and the unmodified maternal genome are passively demethylated at similar rates during the robust cell divisions that occur in the early embryo (69). Even in zygotes, demethylation of both maternal and paternal pronuclei requires DNA replication, a process only partially dependent on Tet3 (14,15). For each observed outcome, the relative contributions of oxi-mC production and DNA demethylation may be distinguished by comparing cells deficient in DNA methyltransferases, which would be depleted for both 5mC and oxi-mC, with cells deficient in TET proteins, which lack only oxi-mC.…”

Section: Discussionmentioning

confidence: 99%

“…In fertilized zygotes, Tet3 was originally thought to oxidize 5mC preferentially in the paternally inherited genome (11)(12)(13); more recently, however, reduced-representation bisulfite sequencing (RRBS) has been used to suggest that demethylation of the maternal genome is also catalyzed by Tet3 (14). RRBS measures the sum of 5mC and 5hmC (vs. C, 5fC, and 5caC) at a fraction of cytosines in the genome, and the data show that loss of 5mC+5hmC in both maternal and paternal pronuclei occurs primarily through a passive, replication-dependent process (14,15). Despite the high expression of Tet3 in oocytes and zygotes (11), Tet3-deficient zygotes display only a marginal increase in 5mC+5hmC in either paternal or maternal pronuclei (14), suggesting the redundant involvement of other TET proteins.…”

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confidence: 99%

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Simultaneous deletion of the methylcytosine oxidases Tet1 and Tet3 increases transcriptome variability in early embryogenesis

Kang

Lienhard

Pastor

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Dioxygenases of the TET (Ten-Eleven Translocation) family produce oxidized methylcytosines, intermediates in DNA demethylation, as well as new epigenetic marks. Here we show data suggesting that TET proteins maintain the consistency of gene transcription. Embryos lacking Tet1 and Tet3 (Tet1/3 DKO) displayed a strong loss of 5-hydroxymethylcytosine (5hmC) and a concurrent increase in 5-methylcytosine (5mC) at the eight-cell stage. Single cells from eight-cell embryos and individual embryonic day 3.5 blastocysts showed unexpectedly variable gene expression compared with controls, and this variability correlated in blastocysts with variably increased 5mC/5hmC in gene bodies and repetitive elements. Despite the variability, genes encoding regulators of cholesterol biosynthesis were reproducibly down-regulated in Tet1/3 DKO blastocysts, resulting in a characteristic phenotype of holoprosencephaly in the few embryos that survived to later stages. Thus, TET enzymes and DNA cytosine modifications could directly or indirectly modulate transcriptional noise, resulting in the selective susceptibility of certain intracellular pathways to regulation by TET proteins.DNA methylation | cholesterol biosynthesis | TET methylcytosine oxidases | 5-hydroxymethylcytosine | 5hmC

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Simultaneous deletion of the methylcytosine oxidases Tet1 and Tet3 increases transcriptome variability in early embryogenesis

Kang

Lienhard

Pastor

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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“…Recent studies have raised the possibility that demethylation can occur through the involvement of the teneleven-translocation family (Tet1, Tet2, and Tet3) that catalyzes the oxidation of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) as a first step in the pathway (1,2). Removal of this unusual base may then be accomplished either by further oxidation followed by base excision repair (3) or through replication dilution (4)(5)(6). Genetic experiments have demonstrated that Tet enzymes are key players during early development, with Tet3-mediated DNA hydroxylation being involved in epigenetic programming of the zygotic paternal DNA (7,8), whereas combinations of Tet1 and Tet2 play a role in the demethylation process that takes place during embryonic stem cell differentiation in vitro (2,(9)(10)(11)(12).…”

mentioning

confidence: 99%

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

Orlanski

Labi

Reizel

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

109

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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.NA methylation takes place at almost all stages of development including the early embryo as well as during lineage commitment and is mediated through a combination of active and passive processes. Recent studies have raised the possibility that demethylation can occur through the involvement of the teneleven-translocation family (Tet1, Tet2, and Tet3) that catalyzes the oxidation of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) as a first step in the pathway (1, 2). Removal of this unusual base may then be accomplished either by further oxidation followed by base excision repair (3) or through replication dilution (4-6). Genetic experiments have demonstrated that Tet enzymes are key players during early development, with Tet3-mediated DNA hydroxylation being involved in epigenetic programming of the zygotic paternal DNA (7, 8), whereas combinations of Tet1 and Tet2 play a role in the demethylation process that takes place during embryonic stem cell differentiation in vitro (2, 9-12).Tet enzymes also contribute to lineage development. Thus, changes in the pattern of 5hmC have been shown to accompany neurogenesis in vivo (13) (21)] appear to alter global 5hmC and 5mC distribution, perturb stem cell self-renewal, cause altered differentiation, and predispose to malignancies (refs. 19, 22, reviewed in ref. 23). None of these studies, however, has addressed the key question of whether demethylation itself is actually required for gene activation and proper lineage differentiation. To this end, we generated a Tet2/Tet3 knockout specific to B-lymphoid development, isolated cells at different stages of differentiation, and analyzed their methylation patterns. Because this approach targets the demethylation machinery in an exclusive manner, it allowed us to evaluate the role of this modification independently of the many transcription factors that drive the process of B-cell differentiation. ResultsIt has already been shown that both Tet2 and Tet3 are highly expressed in B lineage cells (24). With this in mind, we generated Tet2F mice (18, 23) and crossed them with animals expressing Cre under control of the early B-cell-specific Mb1 promoter (25) to obtain mice with a conditional knockout of these enzymes specifically in the B-cell lineage (Materials and Methods). Reduced repres...

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“…However, a number of studies have demonstrated that replicationdependent and -independent (through TET3-mediated oxidation of 5mC to 5hmC) demethylation mechanisms coexist in both male and female pronuclei of mouse zygotes (Salvaing et al 2012, Guo et al 2014a, Shen et al 2014, Wang et al 2014. In the human zygotes at the pronuclear stage, hydroxymethylation of both male and female pronuclei with lower immunofluorescent signal in female pronuclei has been reported by Guo et al (2014b).…”

Section: Discussionmentioning

confidence: 99%

Chromosome hydroxymethylation patterns in human zygotes and cleavage-stage embryos

Efimova¹,

Pendina²,

Tikhonov³

et al. 2015

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We report the sequential changes in 5-hydroxymethylcytosine (5hmC) patterns in the genome of human preimplantation embryos during DNA methylation reprogramming. We have studied chromosome hydroxymethylation and methylation patterns in triploid zygotes and blastomeres of cleavage-stage embryos. Using indirect immunofluorescence, we have analyzed the localization of 5hmC and its co-distribution with 5-methylcytosine (5mC) on the QFH-banded metaphase chromosomes. In zygotes, 5hmC accumulates in both parental chromosome sets, but hydroxymethylation is more intensive in the poorly methylated paternal set. In the maternal set, chromosomes are highly methylated, but contain little 5hmC. Hydroxymethylation is highly region specific in both parental chromosome sets: hydroxymethylated loci correspond to R-bands, but not G-bands, and have well-defined borders, which coincide with the R/G-band boundaries. The centromeric regions and heterochromatin at 1q12, 9q12, 16q11.2, and Yq12 contain little 5mC and no 5hmC. We hypothesize that 5hmC may mark structural/functional genome 'units' corresponding to chromosome bands in the newly formed zygotic genome. In addition, we suggest that the hydroxymethylation of R-bands in zygotes can be treated as a new characteristic distinguishing them from G-bands. At cleavages, chromosomes with asymmetrical hydroxymethylation of sister chromatids appear. They decrease in number during cleavages, whereas totally non-hydroxymethylated chromosomes become numerous. Taken together, our findings suggest that, in the zygotic genome, 5hmC is distributed selectively and its pattern is determined by both parental origin of chromosomes and type of chromosome bands -R, G, or C. At cleavages, chromosome hydroxymethylation pattern is dynamically changed due to passive and non-selective overall loss of 5hmC, which coincides with that of 5mC.

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Active and Passive Demethylation of Male and Female Pronuclear DNA in the Mammalian Zygote

Cited by 329 publications

References 47 publications

Simultaneous deletion of the methylcytosine oxidases Tet1 and Tet3 increases transcriptome variability in early embryogenesis

Simultaneous deletion of the methylcytosine oxidases Tet1 and Tet3 increases transcriptome variability in early embryogenesis

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

Chromosome hydroxymethylation patterns in human zygotes and cleavage-stage embryos

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