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W Reik

Publications and source records attributed to W Reik.

At least 37 records · Page 2Linked to original sources
PubMed2001

Epigenetic targeting in the mouse zygote marks DNA for later methylation: a mechanism for maternal effects in development.

The transgenic sequences in the mouse line TKZ751 are demethylated on a DBA/2 inbred strain background but become highly methylated at postimplantation stages in offspring of a cross with a BALB/c female. In the reciprocal cross the transgene remains demethylated suggesting that imprinted BALB/c methylation modifiers or egg cytoplasmic factors are responsible for this striking maternal effect on de novo methylation. Reciprocal pronuclear transplantation experiments were carried out to distinguish between these mechanisms. The results indicate that a maternally-derived oocyte cytoplasmic factor from BALB/c marks the TKZ751 sequences at fertilization; this mark and postzygotic BALB/c modifiers are both required for de novo methylation of the target sequences at postimplantation stages. Using genetic linkage analyses we mapped the maternal effect to a locus on chromosome 17. Moreover, seven postzygotic modifier loci were identified that increase the postimplantation level of methylation. Analysis of interactions between the maternal and the postzygotic loci shows that both are needed for de novo methylation in the offspring. The combined experiments thus reveal a novel epigenetic marking process at fertilization which targets DNA for later methylation in the foetus. The most significant consequence is that the genotype of the mother can influence the epigenotype of the offspring by this marking process. A number of parental and imprinting effects may be explained by this epigenetic marking.

Alleles↗
PubMed2001

Genomic imprinting: parental influence on the genome.

Genomic imprinting affects several dozen mammalian genes and results in the expression of those genes from only one of the two parental chromosomes. This is brought about by epigenetic instructions--imprints--that are laid down in the parental germ cells. Imprinting is a particularly important genetic mechanism in mammals, and is thought to influence the transfer of nutrients to the fetus and the newborn from the mother. Consistent with this view is the fact that imprinted genes tend to affect growth in the womb and behaviour after birth. Aberrant imprinting disturbs development and is the cause of various disease syndromes. The study of imprinting also provides new insights into epigenetic gene modification during development.

Animals↗
PubMed2001

Culture of preimplantation mouse embryos affects fetal development and the expression of imprinted genes.

Culture of preimplantation mammalian embryos and cells can influence their subsequent growth and differentiation. Previously, we reported that culture of mouse embryonic stem cells is associated with deregulation of genomic imprinting and affects the potential for these cells to develop into normal fetuses. The purpose of our current study was to determine whether culture of preimplantation mouse embryos in a chemically defined medium (M16) with or without fetal calf serum (FCS) can affect their subsequent development and imprinted gene expression. Only one third of the blastocysts that had been cultured from two-cell embryos in M16 medium complemented with FCS developed into viable Day 14 fetuses after transfer into recipients. These M16 + FCS fetuses were reduced in weight as compared with controls and M16 fetuses and had decreased expression of the imprinted H19 and insulin-like growth factor 2 genes associated with a gain of DNA methylation at an imprinting control region upstream of H19. They also displayed increased expression of the imprinted gene Grb10. The growth factor receptor binding gene Grb7, in contrast, was strongly reduced in its expression in most of the M16 + FCS fetuses. No alterations were detected for the imprinted gene MEST: Preimplantation culture in the presence of serum can influence the regulation of multiple growth-related imprinted genes, thus leading to aberrant fetal growth and development.

Animals↗
PubMed2001

A dominant modifier of transgene methylation is mapped by QTL analysis to mouse chromosome 13.

The single-copy hepatitis B virus transgene in the E36 transgenic mouse strain undergoes methylation changes in a parent-of-origin, tissue, and strain-specific fashion. In a C57BL/6 background, the paternally transmitted transgene is methylated in 30% of cells, whereas it is methylated in more than 80% of cells in (BALB/c x C57BL/6) F1 mice. We established previously that several genetic factors were likely to contribute to the transgene methylation profile, some with demethylating and some with de novo methylating activities. Using quantitative trait loci (QTL) mapping, we have now localized one major modifier locus on chromosome 13 (Mod13), which explains a 30% increase in the methylation level of this transgene with no effect on the flanking endogenous sequences. No other QTL could be identified, except for a demethylating activity of low significance located on chromosome 12. Recombinant inbred mice containing a BALB/c allele of Mod13 were then used to show that the presence of Mod13 is sufficient to induce de novo methylation. A segregation between de novo methylation and repression of transgene expression was uncovered, suggesting that this genetic system is also useful for the identification of factors that interpret methylation patterns in the genome.

Animals↗
PubMed2001

Generation of monoparental embryos for investigation into genomic imprinting.

The seminal work of McGrath and Solter (1) and independently of Surani et al. (2) in 1984 established the fundamental principle of nuclear nonequivalency; that is, chromosomes of both paternal and maternal origin are required for development to term in mammals. This was achieved through the creation of diploid reconstituted zygotes, which contained either two maternal or two paternal pronuclei. Embryos containing pronuclei exclusively of maternal or paternal origin display characteristic developmental abnormalities and fail to develop to term. This failure is partially explained by the observation that paternally and maternally derived genomes have complementary roles during embryogenesis, contributing differentially to embryonic and extraembryonic lineages (2-5). These reconstitutions were accomplished by nuclear transplantation and karyoplast fusion using HVJ or Sendai virus-assisted fusion (1). These experiments laid the foundation for the discovery and exploration of this unique form of non-Mendelian mammalian gene regulation whereby expression of genes and hence phenotype were dictated by the parent from whom they where inherited. This parent-of-origin phenomenon is known as genomic imprinting.

Animals↗
PubMed2000

Sequence and functional comparison in the Beckwith-Wiedemann region: implications for a novel imprinting centre and extended imprinting.

The clustered organization of most imprinted genes in mammals suggests coordinated genetic and epigenetic control mechanisms. Comparisons between human and mouse will help in elucidating these mechanisms by identifying structural and functional similarities. Previously we reported on such a comparison in the central part of the mouse imprinting cluster on distal chromosome 7 with the homologous Beckwith-Wiedemann syndrome (BWS) gene cluster on human chromosome 11p15.5. Here we focus on the adjacent sequences of 0.5 Mb including the KCNQ1/Kcnq1 and CDKN1C/Cdkn1c genes, which are implicated in BWS, and on one of the proposed boundary regions of the imprinting cluster. As in the previously analysed central region, this part of the cluster exhibits a highly conserved arrangement and structure of genes. The most striking similarity is found in the 3' part of the KCNQ1/Kcnq1 genes in large stretches of mostly non-coding sequences. The conserved region includes the recently identified KCNQ1OT1/Kcnq1ot1 antisense transcripts, flanked by a strikingly conserved cluster of LINE/Line elements and a CpG island which we show to carry a maternal germline methylation imprint. This region is likely to be the proposed second imprinting centre (IC2) in the BWS cluster. We also identified several novel genes inside and outside the previously proposed boundaries of the imprinting cluster. One of the genes outside the cluster, Obph1, is imprinted in mouse placenta indicating that at least in extra-embryonic tissues the imprinting cluster extends into a larger domain.

Alternative Splicing↗
PubMed2000

Sequence conservation and variability of imprinting in the Beckwith-Wiedemann syndrome gene cluster in human and mouse.

In human and mouse most imprinted genes are arranged in chromosomal clusters. This linked organization suggests coordinated mechanisms controlling imprinted expression. We have sequenced 250 kb in the centre of the mouse imprinting cluster on distal chromosome 7 and compared it with the orthologous Beckwith-Wiedemann gene cluster on human chromosome 11p15.5. This first comparative imprinting cluster analysis revealed a high structural and functional conservation of the six orthologous genes identified. However, several striking differences were also discovered. First, compared with the mouse the human sequence is approximately 40% longer, mostly due to insertions of two large repetitive clusters. One of these clusters encompasses an additional gene coding for a homologue of the ribosomal protein L26. Second, pronounced blocks of unique direct repeats characteristic of imprinted genes were only found in the human sequence. Third, two of the orthologous gene pairs Tssc4/TSSC4 and Ltrpc5/LTRPC5 showed apparent differences in imprinting between human and mouse, whereas others like Tssc6/TSSC6 were not imprinted in either organism. Together these results suggest a significant functional and structural variability in the centre of the imprinting cluster. Some genes escape imprinting in both organisms whereas others exhibit tissue- and species-specific imprinting. Hence the control of imprinting in the cluster appears to be a highly dynamic process under fast evolutionary adaptation. Intriguingly, whereas imprinted genes within the cluster contain CpG islands the non-imprinted Ltrpc5 and Tssc6/TSSC6 do not. This and additional comparisons with other imprinted and non-imprinted regions suggest that CpG islands are key features of imprinted domains.

Amino Acid Sequence↗
PubMed2000

Active demethylation of the paternal genome in the mouse zygote.

DNA methylation is essential for the control of a number of biological mechanisms in mammals [1]. Mammalian development is accompanied by two major waves of genome-wide demethylation and remethylation: one during germ-cell development and the other after fertilisation [2] [3] [4] [5] [6] [7]. Most previous studies have suggested that the genome-wide demethylation observed after fertilisation occurs passively, that is, by the lack of maintenance methylation following DNA replication and cell division [6] [7], although one other study has reported that replication-independent demethylation may also occur during early embryogenesis [8]. Here, we report that genes that are highly methylated in sperm are rapidly demethylated in the zygote only hours after fertilisation, before the first round of DNA replication commences. By contrast, the oocyte-derived maternal alleles are unaffected by this reprogramming. They either remain methylated after fertilisation or become further methylated de novo. These results provide the first direct evidence for active demethylation of single-copy genes in the mammalian zygote and, moreover, reveal a striking asymmetry in epigenetic methylation reprogramming. Whereas paternally (sperm)-derived sequences are exposed to putative active demethylases in the oocyte cytoplasm, maternally (oocyte)-derived sequences are protected from this reaction. These results, whose generality is supported by findings of Mayer et al. [9], have important implications for the establishment of biparental genetic totipotency after fertilisation, the establishment and maintenance of genomic imprinting, and the reprogramming of somatic cells during cloning.

Actins↗
PubMed2000

Deletion of a silencer element in Igf2 results in loss of imprinting independent of H19.

Igf2 and H19 are closely linked, reciprocally imprinted genes on mouse distal chromosome 7. The paternally expressed Igf2 encodes a potent fetal growth factor and the maternally expressed H19 encodes a non-coding RNA (refs 1,2). Shared endoderm-specific enhancers 3' to H19 are necessary for transcription of the maternal copy of H19 and the paternal copy of Igf2 (ref. 3), a chromatin boundary upstream of H19 preventing access of the enhancers to the maternal Igf2 promoters. Mesoderm-specific control elements have not been identified, and the role of differentially methylated regions (DMRs) in Igf2 has not been addressed. Two DMRs in Igf2 are methylated on the active paternal allele, suggesting that they contain silencers. Here we have deleted the DMR1 region in Igf2. Maternal transmission of the deletion results in biallelic expression of Igf2 in most mesodermally derived tissues without altering H19 imprinting or expression. Paternal or maternal transmission leads to continued postnatal transcription of Igf2, in contrast to the wild-type allele, which is silenced soon after birth. These results reveal a mesodermal silencer, which may be regulated by methylation and which has a major role in H19-independent expression and imprinting control of Igf2. Our results establish a new mechanistic principle for imprinted genes whereby epigenetically regulated silencers interact with enhancers to control expression, and suggest a new mechanism for loss of imprinting (LOI) of Igf2, which may be important in a number of diseases.

Animals↗
PubMed2000

Epigenotype-phenotype correlations in Beckwith-Wiedemann syndrome.

Beckwith-Wiedemann syndrome (BWS) is a model imprinting disorder resulting from mutations or epigenetic events involving imprinted genes at chromosome 11p15.5. Thus, germline mutations in CDKN1C, uniparental disomy (UPD), and loss of imprinting of IGF2 and other imprinted genes have been implicated. Many familial BWS cases have germline CDKN1C mutations. However, most BWS cases are sporadic and UPD or putative imprinting errors predominate in this group. We have identified previously a subgroup of sporadic cases with loss of imprinting (LOI) of IGF2 and epigenetic silencing of H19 proposed to be caused by a defect in a distal 11p15.5 imprinting control element (designated BWSIC1). However, many sporadic BWS patients show biallelic IGF2 expression in the presence of normal H19 methylation and expression patterns. This and other evidence suggested the existence of a further imprinting control element (BWSIC2) at 11p15. 5. Recently, we showed that a subgroup of BWS patients have loss of methylation (LOM) at a differentially methylated region (KvDMR1) within the KCNQ1 gene centromeric to the IGF2 and H19 genes. We have now analysed a large series of sporadic cases to define the frequency and phenotypic correlates of epigenetic abnormalities in BWS. LOM at KvDMR1 was detected by Southern analysis or a novel PCR based method in 35 of 69 (51%) sporadic BWS without UPD. LOM at KvDMR1 was often, but not invariably associated with LOI of IGF2. KvDMR1 LOM was not detected in BWS patients with putative BWSIC1 defects and cases with KvDMR1 LOM (that is, putative BWSIC2 defects) invariably had a normal H19 methylation pattern. The incidence of exomphalos in putative BWSIC2 defect patients was not significantly different from that in patients with germline CDKN1C mutations (20/29 and 13/15 respectively), but was significantly greater than that in patients with putative BWSIC1 defects (0/5, p=0.007) and UPD (0/22, p<0.0001). These findings are consistent with the hypothesis that LOM of KvDMR1 (BWSIC2 defect) results in epigenetic silencing of CDKN1C and variable LOI of IGF2. BWS patients with embryonal tumours have UPD or a BWSIC1 defect but not LOM of KvDMR1. This study has further shown how (1) variations in phenotypic expression of BWS may be linked to specific molecular subgroups and (2) molecular analysis of BWS can provide insights into mechanisms of imprinting regulation.

Beckwith-Wiedemann Syndrome↗
PubMed2000

Igf2 imprinting in development and disease.

Igf2 is one of the first imprinted genes discovered and occupies a centre stage in the study of imprinting. This is because it has dramatic effects on the control of fetal growth, it is involved in growth disorders and in cancer, it interacts with products of other imprinted genes, and its imprinting status is under complex regulation in a cluster of tightly linked imprinted genes. Here we review briefly the key features of Igf2 imprinting in normal development and in disease, and hope to show what a fascinating subject of study this gene and its biology provides.

Animals↗
PubMed1999

A maternally methylated CpG island in KvLQT1 is associated with an antisense paternal transcript and loss of imprinting in Beckwith-Wiedemann syndrome.

Loss of imprinting at IGF2, generally through an H19-independent mechanism, is associated with a large percentage of patients with the overgrowth and cancer predisposition condition Beckwith-Wiedemann syndrome (BWS). Imprinting control elements are proposed to exist within the KvLQT1 locus, because multiple BWS-associated chromosome rearrangements disrupt this gene. We have identified an evolutionarily conserved, maternally methylated CpG island (KvDMR1) in an intron of the KvLQT1 gene. Among 12 cases of BWS with normal H19 methylation, 5 showed demethylation of KvDMR1 in fibroblast or lymphocyte DNA; whereas, in 4 cases of BWS with H19 hypermethylation, methylation at KvDMRl was normal. Thus, inactivation of H19 and hypomethylation at KvDMR1 (or an associated phenomenon) represent distinct epigenetic anomalies associated with biallelic expression of IGF2. Reverse transcription-PCR analysis of the human and syntenic mouse loci identified the presence of a KvDMR1-associated RNA transcribed exclusively from the paternal allele and in the opposite orientation with respect to the maternally expressed KvLQT1 gene. We propose that KvDMR1 and/or its associated antisense RNA (KvLQT1-AS) represents an additional imprinting control element or center in the human 11p15.5 and mouse distal 7 imprinted domains.

Animals↗