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CTCF-dependent chromatin bias constitutes transient epigenetic memory of the mother at the H19-Igf2 imprinting control region in prospermatogonia.

Lee DH, Singh P, Tsai SY, Oates N, Spalla A, Spalla C, Brown L, Rivas G, Larson G, Rauch TA, Pfeifer GP, Szabó PE - PLoS Genet. (2010)

Bottom Line: We found that the chromatin composition in fetal germ cells was biased at the ICR between the two alleles with the maternally inherited allele exhibiting more H3K4me3 and less H3K9me3 than the paternally inherited allele.The erasure of these allele-specific chromatin marks is not complete before the process of de novo methylation imprint establishment begins.CTCF-dependent allele-specific chromatin composition imposes a maternal allele-specific delay on de novo methylation imprint establishment at the H19/Igf2 ICR in prospermatogonia.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cellular Biology, Beckman Research Institute, Duarte, CA, USA.

ABSTRACT
Genomic imprints-parental allele-specific DNA methylation marks at the differentially methylated regions (DMRs) of imprinted genes-are erased and reestablished in germ cells according to the individual's sex. Imprint establishment at paternally methylated germ line DMRs occurs in fetal male germ cells. In prospermatogonia, the two unmethylated alleles exhibit different rates of de novo methylation at the H19/Igf2 imprinting control region (ICR) depending on parental origin. We investigated the nature of this epigenetic memory using bisulfite sequencing and allele-specific ChIP-SNuPE assays. We found that the chromatin composition in fetal germ cells was biased at the ICR between the two alleles with the maternally inherited allele exhibiting more H3K4me3 and less H3K9me3 than the paternally inherited allele. We determined genetically that the chromatin bias, and also the delayed methylation establishment in the maternal allele, depended on functional CTCF insulator binding sites in the ICR. Our data suggest that, in primordial germ cells, maternally inherited allele-specific CTCF binding sets up allele-specific chromatin differences at the ICR. The erasure of these allele-specific chromatin marks is not complete before the process of de novo methylation imprint establishment begins. CTCF-dependent allele-specific chromatin composition imposes a maternal allele-specific delay on de novo methylation imprint establishment at the H19/Igf2 ICR in prospermatogonia.

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Model.Functional CTCF sites are required for chromatin bias and delayed methylation of the maternally inherited ICR allele. Expected CTCF binding and chromatin composition is depicted in primordial germ cells (PGC). Observed chromatin bias is depicted in prospermatogonia (PSG). Other details are as Figure 1. The developmental stages are indicated above in dpc. (A) Imprint establishment of the ICR in the normal male germ line. Chromatin bias is observed in the normal ICR between the parental alleles in the absence of CpG methylation at 13.5–14.5 dpc. (B) Imprint establishment at the CTCF site mutant ICR in the male germ line. CTCF cannot bind in the maternal allele in PGCs because of the mutations (x) or in the paternal allele because of CpG methylation. The chromatin bias, found in normal cells, is no longer observed between parental alleles in the mutant cells at 13.5–14.5 dpc and the maternal allele's methylation is not delayed at 15.5–17.5 dpc.
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pgen-1001224-g007: Model.Functional CTCF sites are required for chromatin bias and delayed methylation of the maternally inherited ICR allele. Expected CTCF binding and chromatin composition is depicted in primordial germ cells (PGC). Observed chromatin bias is depicted in prospermatogonia (PSG). Other details are as Figure 1. The developmental stages are indicated above in dpc. (A) Imprint establishment of the ICR in the normal male germ line. Chromatin bias is observed in the normal ICR between the parental alleles in the absence of CpG methylation at 13.5–14.5 dpc. (B) Imprint establishment at the CTCF site mutant ICR in the male germ line. CTCF cannot bind in the maternal allele in PGCs because of the mutations (x) or in the paternal allele because of CpG methylation. The chromatin bias, found in normal cells, is no longer observed between parental alleles in the mutant cells at 13.5–14.5 dpc and the maternal allele's methylation is not delayed at 15.5–17.5 dpc.

Mentions: We found a slight (∼10%), but reproducible bias in the H3K4me2 levels toward the maternally inherited allele in male and female germ cell ChIP samples at 13.5 and 14.5 dpc (Figure 5). The bias was present in the ICR at −3 kb and −4 kb positions. H3K4me2 enrichment in germ cells was similar to the level found in MEFs (Figure S9), suggesting that the ICR had not been stripped of this mark at 13.5–14.5 dpc. H3K9me3 was reciprocally biased: the paternally inherited allele exhibited about 10% higher enrichment at 13.5 and 14.5 dpc (Figure 6). The allele-specificity of the bias for H3K4me2 and H3K9me3 in 13.5 dpc germ cells was in agreement with the somatic pattern (Figure S6A), being maternal and paternal specific, respectively, suggesting that it originates in premigratory PGCs (Figure 7A). The amplitude of the bias, however, was smaller than in the soma, consistent with the possibility that the chromatin differences are being erased in germ cells around mid-gestation and only the remnants of the allele-specific differences can be detected at 13.5–14.5 dpc. H3K9me3 levels at the ICR, however, were very low in germ cells at these stages (not shown), consistent with the possibility that similarly to CTCF but unlike H3K4me2 this mark is almost completely removed by 13.5 dpc.


CTCF-dependent chromatin bias constitutes transient epigenetic memory of the mother at the H19-Igf2 imprinting control region in prospermatogonia.

Lee DH, Singh P, Tsai SY, Oates N, Spalla A, Spalla C, Brown L, Rivas G, Larson G, Rauch TA, Pfeifer GP, Szabó PE - PLoS Genet. (2010)

Model.Functional CTCF sites are required for chromatin bias and delayed methylation of the maternally inherited ICR allele. Expected CTCF binding and chromatin composition is depicted in primordial germ cells (PGC). Observed chromatin bias is depicted in prospermatogonia (PSG). Other details are as Figure 1. The developmental stages are indicated above in dpc. (A) Imprint establishment of the ICR in the normal male germ line. Chromatin bias is observed in the normal ICR between the parental alleles in the absence of CpG methylation at 13.5–14.5 dpc. (B) Imprint establishment at the CTCF site mutant ICR in the male germ line. CTCF cannot bind in the maternal allele in PGCs because of the mutations (x) or in the paternal allele because of CpG methylation. The chromatin bias, found in normal cells, is no longer observed between parental alleles in the mutant cells at 13.5–14.5 dpc and the maternal allele's methylation is not delayed at 15.5–17.5 dpc.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2991272&req=5

pgen-1001224-g007: Model.Functional CTCF sites are required for chromatin bias and delayed methylation of the maternally inherited ICR allele. Expected CTCF binding and chromatin composition is depicted in primordial germ cells (PGC). Observed chromatin bias is depicted in prospermatogonia (PSG). Other details are as Figure 1. The developmental stages are indicated above in dpc. (A) Imprint establishment of the ICR in the normal male germ line. Chromatin bias is observed in the normal ICR between the parental alleles in the absence of CpG methylation at 13.5–14.5 dpc. (B) Imprint establishment at the CTCF site mutant ICR in the male germ line. CTCF cannot bind in the maternal allele in PGCs because of the mutations (x) or in the paternal allele because of CpG methylation. The chromatin bias, found in normal cells, is no longer observed between parental alleles in the mutant cells at 13.5–14.5 dpc and the maternal allele's methylation is not delayed at 15.5–17.5 dpc.
Mentions: We found a slight (∼10%), but reproducible bias in the H3K4me2 levels toward the maternally inherited allele in male and female germ cell ChIP samples at 13.5 and 14.5 dpc (Figure 5). The bias was present in the ICR at −3 kb and −4 kb positions. H3K4me2 enrichment in germ cells was similar to the level found in MEFs (Figure S9), suggesting that the ICR had not been stripped of this mark at 13.5–14.5 dpc. H3K9me3 was reciprocally biased: the paternally inherited allele exhibited about 10% higher enrichment at 13.5 and 14.5 dpc (Figure 6). The allele-specificity of the bias for H3K4me2 and H3K9me3 in 13.5 dpc germ cells was in agreement with the somatic pattern (Figure S6A), being maternal and paternal specific, respectively, suggesting that it originates in premigratory PGCs (Figure 7A). The amplitude of the bias, however, was smaller than in the soma, consistent with the possibility that the chromatin differences are being erased in germ cells around mid-gestation and only the remnants of the allele-specific differences can be detected at 13.5–14.5 dpc. H3K9me3 levels at the ICR, however, were very low in germ cells at these stages (not shown), consistent with the possibility that similarly to CTCF but unlike H3K4me2 this mark is almost completely removed by 13.5 dpc.

Bottom Line: We found that the chromatin composition in fetal germ cells was biased at the ICR between the two alleles with the maternally inherited allele exhibiting more H3K4me3 and less H3K9me3 than the paternally inherited allele.The erasure of these allele-specific chromatin marks is not complete before the process of de novo methylation imprint establishment begins.CTCF-dependent allele-specific chromatin composition imposes a maternal allele-specific delay on de novo methylation imprint establishment at the H19/Igf2 ICR in prospermatogonia.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cellular Biology, Beckman Research Institute, Duarte, CA, USA.

ABSTRACT
Genomic imprints-parental allele-specific DNA methylation marks at the differentially methylated regions (DMRs) of imprinted genes-are erased and reestablished in germ cells according to the individual's sex. Imprint establishment at paternally methylated germ line DMRs occurs in fetal male germ cells. In prospermatogonia, the two unmethylated alleles exhibit different rates of de novo methylation at the H19/Igf2 imprinting control region (ICR) depending on parental origin. We investigated the nature of this epigenetic memory using bisulfite sequencing and allele-specific ChIP-SNuPE assays. We found that the chromatin composition in fetal germ cells was biased at the ICR between the two alleles with the maternally inherited allele exhibiting more H3K4me3 and less H3K9me3 than the paternally inherited allele. We determined genetically that the chromatin bias, and also the delayed methylation establishment in the maternal allele, depended on functional CTCF insulator binding sites in the ICR. Our data suggest that, in primordial germ cells, maternally inherited allele-specific CTCF binding sets up allele-specific chromatin differences at the ICR. The erasure of these allele-specific chromatin marks is not complete before the process of de novo methylation imprint establishment begins. CTCF-dependent allele-specific chromatin composition imposes a maternal allele-specific delay on de novo methylation imprint establishment at the H19/Igf2 ICR in prospermatogonia.

Show MeSH
Related in: MedlinePlus