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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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The imprint cycle at the H19/Igf2 ICR.Schematic representation of epigenetic features at the H19/Igf2 imprinted domain based on publications referenced in the Introduction. (A) The H19/Igf2 imprinted domain in the soma. Maternal chromosome (M): unmethylated (white lollipops) ICR (shaded area) is inherited from the egg. CTCF protein (yellow ovals) at binding sites 1–2 and 3–4 at about −4 kb and −3 kb upstream of the H19 transcription start site imparts insulator activity (bracket) between the Igf2 promoters and the shared, downstream enhancers (orange oval). Paternal chromosome (P): methylated (black lollipops) ICR is inherited from the sperm, CTCF cannot bind, hence ICR has no insulator activity, Igf2 promoters and enhancers can interact. Early in postimplantation development, the H19 promoter is inactivated by an ICR-dependent mechanism (horizontal arrow). Active or repressive chromatin (green or red hexagon) is present at expressed or silent alleles of genes (green-red rectangles) and at respective alleles of the ICR. (B) Fate of the imprint in the female germ line. Methylation status of the ICR is depicted in the mature oocyte (OC), spermatozoon (SPZ), primordial germ cells (PGC) primary oocytes (POC) at gestational stages (in dpc). (C) Fate of the imprint in the male germ line. Methylation status is depicted in OC, SPZ and PGC as above and in prospermatogonia (PSG), spermatogonia (SG) pachytene spermatocytes (PS) and round spermatids (ST). The developmental stage under investigation is marked by a rectangle.
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pgen-1001224-g001: The imprint cycle at the H19/Igf2 ICR.Schematic representation of epigenetic features at the H19/Igf2 imprinted domain based on publications referenced in the Introduction. (A) The H19/Igf2 imprinted domain in the soma. Maternal chromosome (M): unmethylated (white lollipops) ICR (shaded area) is inherited from the egg. CTCF protein (yellow ovals) at binding sites 1–2 and 3–4 at about −4 kb and −3 kb upstream of the H19 transcription start site imparts insulator activity (bracket) between the Igf2 promoters and the shared, downstream enhancers (orange oval). Paternal chromosome (P): methylated (black lollipops) ICR is inherited from the sperm, CTCF cannot bind, hence ICR has no insulator activity, Igf2 promoters and enhancers can interact. Early in postimplantation development, the H19 promoter is inactivated by an ICR-dependent mechanism (horizontal arrow). Active or repressive chromatin (green or red hexagon) is present at expressed or silent alleles of genes (green-red rectangles) and at respective alleles of the ICR. (B) Fate of the imprint in the female germ line. Methylation status of the ICR is depicted in the mature oocyte (OC), spermatozoon (SPZ), primordial germ cells (PGC) primary oocytes (POC) at gestational stages (in dpc). (C) Fate of the imprint in the male germ line. Methylation status is depicted in OC, SPZ and PGC as above and in prospermatogonia (PSG), spermatogonia (SG) pachytene spermatocytes (PS) and round spermatids (ST). The developmental stage under investigation is marked by a rectangle.

Mentions: The paternally expressed insulin-like growth factor 2 (Igf2) and maternally expressed H19 genes on mouse distal chromosome 7 [13] are coordinately expressed during embryonic development, due to shared tissue-specific enhancers (Figure 1A) [14], [15]. A paternally methylated germ line DMR between Igf2 and H19 [16]–[18] is responsible for monoallelic expression of both H19 and Igf2 [19]–[21], and therefore, is called an imprinting control region (ICR). The regulatory functions of the ICR depend on allele-specific DNA methylation. Inactivation of the H19 promoter takes place in post-implantation development on the paternal chromosome and it depends on ICR methylation [22]. The ICR functions as a methylation regulated enhancer blocker [23]–[27]: CTCF protein [28]–[30] binds in the unmethylated maternal allele and insulates between the Igf2 promoters and the shared enhancers. DNA methylation in the paternal allele inhibits CTCF binding, hence the ICR has no insulator activity, and the Igf2 promoters and the enhancers can interact. Targeted mutagenesis of the CTCF binding sites in the mouse results in a loss of enhancer-blocking activity and increased DNA methylation in the mutant maternal chromosome [31]–[33]. CTCF binding in the ICR is the major organizer of chromatin composition in the maternal allele along the entire imprinted domain [34]–[36]. CTCF recruits active histone tail modification marks to the ICR and to the H19 gene [34] and also recruits at a distance, Polycomb-mediated H3K27me3 repressive marks at the Igf2 promoter and at the Igf2 DMRs [34], [35].


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)

The imprint cycle at the H19/Igf2 ICR.Schematic representation of epigenetic features at the H19/Igf2 imprinted domain based on publications referenced in the Introduction. (A) The H19/Igf2 imprinted domain in the soma. Maternal chromosome (M): unmethylated (white lollipops) ICR (shaded area) is inherited from the egg. CTCF protein (yellow ovals) at binding sites 1–2 and 3–4 at about −4 kb and −3 kb upstream of the H19 transcription start site imparts insulator activity (bracket) between the Igf2 promoters and the shared, downstream enhancers (orange oval). Paternal chromosome (P): methylated (black lollipops) ICR is inherited from the sperm, CTCF cannot bind, hence ICR has no insulator activity, Igf2 promoters and enhancers can interact. Early in postimplantation development, the H19 promoter is inactivated by an ICR-dependent mechanism (horizontal arrow). Active or repressive chromatin (green or red hexagon) is present at expressed or silent alleles of genes (green-red rectangles) and at respective alleles of the ICR. (B) Fate of the imprint in the female germ line. Methylation status of the ICR is depicted in the mature oocyte (OC), spermatozoon (SPZ), primordial germ cells (PGC) primary oocytes (POC) at gestational stages (in dpc). (C) Fate of the imprint in the male germ line. Methylation status is depicted in OC, SPZ and PGC as above and in prospermatogonia (PSG), spermatogonia (SG) pachytene spermatocytes (PS) and round spermatids (ST). The developmental stage under investigation is marked by a rectangle.
© Copyright Policy
Related In: Results  -  Collection

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

pgen-1001224-g001: The imprint cycle at the H19/Igf2 ICR.Schematic representation of epigenetic features at the H19/Igf2 imprinted domain based on publications referenced in the Introduction. (A) The H19/Igf2 imprinted domain in the soma. Maternal chromosome (M): unmethylated (white lollipops) ICR (shaded area) is inherited from the egg. CTCF protein (yellow ovals) at binding sites 1–2 and 3–4 at about −4 kb and −3 kb upstream of the H19 transcription start site imparts insulator activity (bracket) between the Igf2 promoters and the shared, downstream enhancers (orange oval). Paternal chromosome (P): methylated (black lollipops) ICR is inherited from the sperm, CTCF cannot bind, hence ICR has no insulator activity, Igf2 promoters and enhancers can interact. Early in postimplantation development, the H19 promoter is inactivated by an ICR-dependent mechanism (horizontal arrow). Active or repressive chromatin (green or red hexagon) is present at expressed or silent alleles of genes (green-red rectangles) and at respective alleles of the ICR. (B) Fate of the imprint in the female germ line. Methylation status of the ICR is depicted in the mature oocyte (OC), spermatozoon (SPZ), primordial germ cells (PGC) primary oocytes (POC) at gestational stages (in dpc). (C) Fate of the imprint in the male germ line. Methylation status is depicted in OC, SPZ and PGC as above and in prospermatogonia (PSG), spermatogonia (SG) pachytene spermatocytes (PS) and round spermatids (ST). The developmental stage under investigation is marked by a rectangle.
Mentions: The paternally expressed insulin-like growth factor 2 (Igf2) and maternally expressed H19 genes on mouse distal chromosome 7 [13] are coordinately expressed during embryonic development, due to shared tissue-specific enhancers (Figure 1A) [14], [15]. A paternally methylated germ line DMR between Igf2 and H19 [16]–[18] is responsible for monoallelic expression of both H19 and Igf2 [19]–[21], and therefore, is called an imprinting control region (ICR). The regulatory functions of the ICR depend on allele-specific DNA methylation. Inactivation of the H19 promoter takes place in post-implantation development on the paternal chromosome and it depends on ICR methylation [22]. The ICR functions as a methylation regulated enhancer blocker [23]–[27]: CTCF protein [28]–[30] binds in the unmethylated maternal allele and insulates between the Igf2 promoters and the shared enhancers. DNA methylation in the paternal allele inhibits CTCF binding, hence the ICR has no insulator activity, and the Igf2 promoters and the enhancers can interact. Targeted mutagenesis of the CTCF binding sites in the mouse results in a loss of enhancer-blocking activity and increased DNA methylation in the mutant maternal chromosome [31]–[33]. CTCF binding in the ICR is the major organizer of chromatin composition in the maternal allele along the entire imprinted domain [34]–[36]. CTCF recruits active histone tail modification marks to the ICR and to the H19 gene [34] and also recruits at a distance, Polycomb-mediated H3K27me3 repressive marks at the Igf2 promoter and at the Igf2 DMRs [34], [35].

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