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Absence of Maternal Methylation in Biparental Hydatidiform Moles from Women with NLRP7 Maternal-Effect Mutations Reveals Widespread Placenta-Specific Imprinting.

Sanchez-Delgado M, Martin-Trujillo A, Tayama C, Vidal E, Esteller M, Iglesias-Platas I, Deo N, Barney O, Maclean K, Hata K, Nakabayashi K, Fisher R, Monk D - PLoS Genet. (2015)

Bottom Line: These epigenotypes are consistent with NLRP7 being a maternal-effect gene and involved in imprint acquisition in the oocyte.In addition, bioinformatic screening of the resulting methylation datasets identified over sixty loci with methylation profiles consistent with imprinting in the placenta, of which we confirm 22 as novel maternally methylated loci.These observations strongly suggest that the molar phenotypes are due to defective placenta-specific imprinting and over-expression of paternally expressed transcripts, highlighting that maternal-effect mutations of NLRP7 are associated with the most severe form of multi-locus imprinting defects in humans.

View Article: PubMed Central - PubMed

Affiliation: Imprinting and Cancer Group, Cancer Epigenetic and Biology Program, Institut d'Investigació Biomedica de Bellvitge, Hospital Duran i Reynals, Barcelona, Spain.

ABSTRACT
Familial recurrent hydatidiform mole (RHM) is a maternal-effect autosomal recessive disorder usually associated with mutations of the NLRP7 gene. It is characterized by HM with excessive trophoblastic proliferation, which mimics the appearance of androgenetic molar conceptuses despite their diploid biparental constitution. It has been proposed that the phenotypes of both types of mole are associated with aberrant genomic imprinting. However no systematic analyses for imprinting defects have been reported. Here, we present the genome-wide methylation profiles of both spontaneous androgenetic and biparental NLRP7 defective molar tissues. We observe total paternalization of all ubiquitous and placenta-specific differentially methylated regions (DMRs) in four androgenetic moles; namely gain of methylation at paternally methylated loci and absence of methylation at maternally methylated regions. The methylation defects observed in five RHM biopsies from NLRP7 defective patients are restricted to lack-of-methylation at maternal DMRs. Surprisingly RHMs from two sisters with the same missense mutations, as well as consecutive RHMs from one affected female show subtle allelic methylation differences, suggesting inter-RHM variation. These epigenotypes are consistent with NLRP7 being a maternal-effect gene and involved in imprint acquisition in the oocyte. In addition, bioinformatic screening of the resulting methylation datasets identified over sixty loci with methylation profiles consistent with imprinting in the placenta, of which we confirm 22 as novel maternally methylated loci. These observations strongly suggest that the molar phenotypes are due to defective placenta-specific imprinting and over-expression of paternally expressed transcripts, highlighting that maternal-effect mutations of NLRP7 are associated with the most severe form of multi-locus imprinting defects in humans.

No MeSH data available.


Related in: MedlinePlus

Allele-specific RT-PCR analysis of candidate placenta-specific imprinted genes.(A) Confirmation of paternal expression of RHOBTB3, SCIN and ZNF396 in term placenta and biallelic expression of neighboring genes. (B) The allele-specific expression analysis of genes flanking known placenta-specific imprinted transcripts GPR1-AS, MCCC1 and AGBL3. Biallelic expression of ZNF396, ADAM23, MCCC1 and AGBL3 was confirmed in somatic tissues. (C) Allele-specific RT-PCR analysis of Adam23 in mouse embryo and placenta at embryonic day 9.5. The asterisk (*) in the sequence traces shows the position of the polymorphic base. The blue boxes in the figures represent the paternally expressed transcripts, white boxes signify biallelically expressed genes and grey boxes are transcripts not expressed in term placenta samples. The location of unmethylated CpG islands and the DMRs are shown by the lollipops. PL = placenta, BR = brain, KID = kidney, LY = blood leucocytes, CB = cord blood.
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pgen.1005644.g004: Allele-specific RT-PCR analysis of candidate placenta-specific imprinted genes.(A) Confirmation of paternal expression of RHOBTB3, SCIN and ZNF396 in term placenta and biallelic expression of neighboring genes. (B) The allele-specific expression analysis of genes flanking known placenta-specific imprinted transcripts GPR1-AS, MCCC1 and AGBL3. Biallelic expression of ZNF396, ADAM23, MCCC1 and AGBL3 was confirmed in somatic tissues. (C) Allele-specific RT-PCR analysis of Adam23 in mouse embryo and placenta at embryonic day 9.5. The asterisk (*) in the sequence traces shows the position of the polymorphic base. The blue boxes in the figures represent the paternally expressed transcripts, white boxes signify biallelically expressed genes and grey boxes are transcripts not expressed in term placenta samples. The location of unmethylated CpG islands and the DMRs are shown by the lollipops. PL = placenta, BR = brain, KID = kidney, LY = blood leucocytes, CB = cord blood.

Mentions: The main biological significance of allele-specific methylation is allele-specific RNA expression, which in the case of maternally methylated regions is predicted to dictate paternal expression. We subsequently determined allelic expression for a subset of transcripts that contained highly polymorphic exonic SNPs. Allele-specific RT-PCR confirmed paternal expression of RHOBTB3, SCIN, ZNF396, ST8SIA1, ZFP90, CCDC71L, RASGRF1, HECW1 and CMTM3 with monoallelic expression of CD83 in a polymorphic fashion in multiple placental biopsies (Fig 4A and S6A Fig; S3 Table). In situations where monoallelic expression was uninformative due to maternal DNA also being heterozygous, the methylated allele was always the repressed one, suggesting a functional link between methylation and expression. In addition, we identified a maternally methylated CpG islands overlapping the promoter of SNCB, a transcript that has previously been described as paternally expressed in placenta [21]. Furthermore, we also identified a maternal DMR within the TTC39A gene that is adjacent to EPS15, a transcript also reported to be imprinted in placenta [22] (S5 Fig).


Absence of Maternal Methylation in Biparental Hydatidiform Moles from Women with NLRP7 Maternal-Effect Mutations Reveals Widespread Placenta-Specific Imprinting.

Sanchez-Delgado M, Martin-Trujillo A, Tayama C, Vidal E, Esteller M, Iglesias-Platas I, Deo N, Barney O, Maclean K, Hata K, Nakabayashi K, Fisher R, Monk D - PLoS Genet. (2015)

Allele-specific RT-PCR analysis of candidate placenta-specific imprinted genes.(A) Confirmation of paternal expression of RHOBTB3, SCIN and ZNF396 in term placenta and biallelic expression of neighboring genes. (B) The allele-specific expression analysis of genes flanking known placenta-specific imprinted transcripts GPR1-AS, MCCC1 and AGBL3. Biallelic expression of ZNF396, ADAM23, MCCC1 and AGBL3 was confirmed in somatic tissues. (C) Allele-specific RT-PCR analysis of Adam23 in mouse embryo and placenta at embryonic day 9.5. The asterisk (*) in the sequence traces shows the position of the polymorphic base. The blue boxes in the figures represent the paternally expressed transcripts, white boxes signify biallelically expressed genes and grey boxes are transcripts not expressed in term placenta samples. The location of unmethylated CpG islands and the DMRs are shown by the lollipops. PL = placenta, BR = brain, KID = kidney, LY = blood leucocytes, CB = cord blood.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4636177&req=5

pgen.1005644.g004: Allele-specific RT-PCR analysis of candidate placenta-specific imprinted genes.(A) Confirmation of paternal expression of RHOBTB3, SCIN and ZNF396 in term placenta and biallelic expression of neighboring genes. (B) The allele-specific expression analysis of genes flanking known placenta-specific imprinted transcripts GPR1-AS, MCCC1 and AGBL3. Biallelic expression of ZNF396, ADAM23, MCCC1 and AGBL3 was confirmed in somatic tissues. (C) Allele-specific RT-PCR analysis of Adam23 in mouse embryo and placenta at embryonic day 9.5. The asterisk (*) in the sequence traces shows the position of the polymorphic base. The blue boxes in the figures represent the paternally expressed transcripts, white boxes signify biallelically expressed genes and grey boxes are transcripts not expressed in term placenta samples. The location of unmethylated CpG islands and the DMRs are shown by the lollipops. PL = placenta, BR = brain, KID = kidney, LY = blood leucocytes, CB = cord blood.
Mentions: The main biological significance of allele-specific methylation is allele-specific RNA expression, which in the case of maternally methylated regions is predicted to dictate paternal expression. We subsequently determined allelic expression for a subset of transcripts that contained highly polymorphic exonic SNPs. Allele-specific RT-PCR confirmed paternal expression of RHOBTB3, SCIN, ZNF396, ST8SIA1, ZFP90, CCDC71L, RASGRF1, HECW1 and CMTM3 with monoallelic expression of CD83 in a polymorphic fashion in multiple placental biopsies (Fig 4A and S6A Fig; S3 Table). In situations where monoallelic expression was uninformative due to maternal DNA also being heterozygous, the methylated allele was always the repressed one, suggesting a functional link between methylation and expression. In addition, we identified a maternally methylated CpG islands overlapping the promoter of SNCB, a transcript that has previously been described as paternally expressed in placenta [21]. Furthermore, we also identified a maternal DMR within the TTC39A gene that is adjacent to EPS15, a transcript also reported to be imprinted in placenta [22] (S5 Fig).

Bottom Line: These epigenotypes are consistent with NLRP7 being a maternal-effect gene and involved in imprint acquisition in the oocyte.In addition, bioinformatic screening of the resulting methylation datasets identified over sixty loci with methylation profiles consistent with imprinting in the placenta, of which we confirm 22 as novel maternally methylated loci.These observations strongly suggest that the molar phenotypes are due to defective placenta-specific imprinting and over-expression of paternally expressed transcripts, highlighting that maternal-effect mutations of NLRP7 are associated with the most severe form of multi-locus imprinting defects in humans.

View Article: PubMed Central - PubMed

Affiliation: Imprinting and Cancer Group, Cancer Epigenetic and Biology Program, Institut d'Investigació Biomedica de Bellvitge, Hospital Duran i Reynals, Barcelona, Spain.

ABSTRACT
Familial recurrent hydatidiform mole (RHM) is a maternal-effect autosomal recessive disorder usually associated with mutations of the NLRP7 gene. It is characterized by HM with excessive trophoblastic proliferation, which mimics the appearance of androgenetic molar conceptuses despite their diploid biparental constitution. It has been proposed that the phenotypes of both types of mole are associated with aberrant genomic imprinting. However no systematic analyses for imprinting defects have been reported. Here, we present the genome-wide methylation profiles of both spontaneous androgenetic and biparental NLRP7 defective molar tissues. We observe total paternalization of all ubiquitous and placenta-specific differentially methylated regions (DMRs) in four androgenetic moles; namely gain of methylation at paternally methylated loci and absence of methylation at maternally methylated regions. The methylation defects observed in five RHM biopsies from NLRP7 defective patients are restricted to lack-of-methylation at maternal DMRs. Surprisingly RHMs from two sisters with the same missense mutations, as well as consecutive RHMs from one affected female show subtle allelic methylation differences, suggesting inter-RHM variation. These epigenotypes are consistent with NLRP7 being a maternal-effect gene and involved in imprint acquisition in the oocyte. In addition, bioinformatic screening of the resulting methylation datasets identified over sixty loci with methylation profiles consistent with imprinting in the placenta, of which we confirm 22 as novel maternally methylated loci. These observations strongly suggest that the molar phenotypes are due to defective placenta-specific imprinting and over-expression of paternally expressed transcripts, highlighting that maternal-effect mutations of NLRP7 are associated with the most severe form of multi-locus imprinting defects in humans.

No MeSH data available.


Related in: MedlinePlus