Paternally Expressed Genes Research Articles

The zinc finger transcription factor PW1/PEG3 restrains murine beta cell cycling.

Aims/hypothesisPw1 or paternally-expressed gene 3 (Peg3) encodes a zinc finger transcription factor that is widely expressed during mouse embryonic development and later restricted to multiple somatic stem cell lineages in the adult. The aim of the present study was to define Pw1 expression in the embryonic and adult pancreas and investigate its role in the beta cell cycle in Pw1 wild-type and mutant mice.MethodsWe analysed PW1 expression by immunohistochemistry in pancreas of nonpregant and pregnant mice and following injury by partial duct ligation. Its role in the beta cell cycle was studied in vivo using a novel conditional knockout mouse and in vitro by lentivirus-mediated gene knockdown.ResultsWe showed that PW1 is expressed in early pancreatic progenitors at E9.5 but becomes progressively restricted to fully differentiated beta cells as they become established after birth and withdraw from the cell cycle. Notably, PW1 expression declines when beta cells are induced to proliferate and loss of PW1 function activates the beta cell cycle.Conclusions/interpretationThese results indicate that PW1 is a co-regulator of the beta cell cycle and can thus be considered a novel therapeutic target in diabetes.Electronic supplementary materialThe online version of this article (doi:10.1007/s00125-016-3954-z) contains peer-reviewed but unedited supplementary material, which is available to authorised users.

An Epigenetic Role for Disrupted Paternal Gene Expression in Postzygotic Seed Abortion in Arabidopsis Interspecific Hybrids

Interspecific hybrids often increase the levels of heterozygosity and hybrid vigor, but some interspecific hybrid seeds are aborted shortly after fertilization. The mechanism behind this postzygotic seed abortion is poorly understood. Here, we report genome-wide analysis of allelic expression changes in developing siliques and seeds in three F1 interspecific crosses between Arabidopsis thaliana (Col, Ler, or C24) and Arabidopsis arenosa. The majority of maternally expressed genes (MEGs) were shared among all three F1 interspecific crosses, whereas ∼90% of 272 paternally expressed genes (PEGs) were found only in one or two F1 crosses, suggesting a role for disrupted paternal gene expression in seed abortion that varies in different crosses. Consistent with this notion, 12 PEGs in the infertile interspecific hybrids matched MEGs in fertile intraspecific hybrids. This disruption of PEGs in the interspecific hybrids was consistent with the upregulation of the genes in the paternal-excess interploidy cross (2X6) between a diploid mother and a hexaploid father, leading to the seed abortion. Moreover, a subset of PEGs in the interspecific crosses were also upregulated in the intraspecific hybrid met1XWT or meaXWT, in which the mutant of MET1 (DNA METHYLTRANSFERASE1) or MEDEA, a Polycomb Repressive Complex2 gene, was used as the maternal parent. These data suggest that maternal epigenetic factors and paternal gene expression play important roles in the postzygotic seed abortion in interspecific hybrids or neo-allopolyploids.

Influence of the Prader-Willi syndrome imprinting center on the DNA methylation landscape in the mouse brain

Reduced representation bisulfite sequencing (RRBS) was used to analyze DNA methylation patterns across the mouse brain genome in mice carrying a deletion of the Prader-Willi syndrome imprinting center (PWS-IC) on either the maternally- or paternally-inherited chromosome. Within the ∼3.7 Mb imprinted Angelman/Prader-Willi syndrome (AS/PWS) domain, 254 CpG sites were interrogated for changes in methylation due to PWS-IC deletion. Paternally-inherited deletion of the PWS-IC increased methylation levels ∼2-fold at each CpG site (compared to wild-type controls) at differentially methylated regions (DMRs) associated with 5′ CpG island promoters of paternally-expressed genes; these methylation changes extended, to a variable degree, into the adjacent CpG island shores. Maternal PWS-IC deletion yielded little or no changes in methylation at these DMRs, and methylation of CpG sites outside of promoter DMRs also was unchanged upon maternal or paternal PWS-IC deletion. Using stringent ascertainment criteria, ∼750,000 additional CpG sites were also interrogated across the entire mouse genome. This analysis identified 26 loci outside of the imprinted AS/PWS domain showing altered DNA methylation levels of ≥25% upon PWS-IC deletion. Curiously, altered methylation at 9 of these loci was a consequence of maternal PWS-IC deletion (maternal PWS-IC deletion by itself is not known to be associated with a phenotype in either humans or mice), and 10 of these loci exhibited the same changes in methylation irrespective of the parental origin of the PWS-IC deletion. These results suggest that the PWS-IC may affect DNA methylation at these loci by directly interacting with them, or may affect methylation at these loci through indirect downstream effects due to PWS-IC deletion. They further suggest the PWS-IC may have a previously uncharacterized function outside of the imprinted AS/PWS domain.

TSG101 and PEG10 are prognostic markers in squamous cell/adenosquamous carcinomas and adenocarcinoma of the gallbladder.

The clinicopathological characteristics of squamous cell/adenosquamous carcinoma (SC/ASC) are currently not well documented, and as the prevalence of SC/ASC is uncommon in gallbladder cancers, a prognostic marker has not yet been found. In the present study, the expression of tumor susceptibility gene (TSG) 101 and paternally expressed gene (PEG) 10 was assessed in 46 SC/ASCs and 80 adenocarcinomas (ACs) using immunohistochemistry, and the samples were further analyzed to examine correlations with the clinicopathological characteristics. It was demonstrated that positive TSG101 and PEG10 expression were significantly associated with large tumor size, high tumor-node-metastasis (TNM) stage, lymph node metastasis, invasion and no resection (only biopsy) of SC/ASC and AC. The univariate Kaplan-Meier analysis showed that positive TSG101 and PEG10 expression, and differentiation, tumor size, TNM stage, lymph node metastasis, invasion and surgical curability, is closely associated with a decreased overall survival in SC/ASC and AC patients (P<0.05 or P<0.001). The multivariate Cox regression analysis identified that positive TSG101 and PEG10 expression are independent factors for a poor-prognosis in SC/ASC and AC patients. The present study indicates that positive TSG101 and PEG10 expression are closely associated with the clinical, pathological and biological behaviors, and a poor prognosis in gallbladder cancer.

Genomic Imprinting in the Arabidopsis Embryo Is Partly Regulated by PRC2

Genomic imprinting results in monoallelic gene expression in a parent-of-origin-dependent manner and is regulated by the differential epigenetic marking of the parental alleles. In plants, genomic imprinting has been primarily described for genes expressed in the endosperm, a tissue nourishing the developing embryo that does not contribute to the next generation. In Arabidopsis, the genes MEDEA (MEA) and PHERES1 (PHE1), which are imprinted in the endosperm, are also expressed in the embryo; whether their embryonic expression is regulated by imprinting or not, however, remains controversial. In contrast, the maternally expressed in embryo 1 (mee1) gene of maize is clearly imprinted in the embryo. We identified several imprinted candidate genes in an allele-specific transcriptome of hybrid Arabidopsis embryos and confirmed parent-of-origin-dependent, monoallelic expression for eleven maternally expressed genes (MEGs) and one paternally expressed gene (PEG) in the embryo, using allele-specific expression analyses and reporter gene assays. Genetic studies indicate that the Polycomb Repressive Complex 2 (PRC2) but not the DNA METHYLTRANSFERASE1 (MET1) is involved in regulating imprinted expression in the embryo. In the seedling, all embryonic MEGs and the PEG are expressed from both parents, suggesting that the imprint is erased during late embryogenesis or early vegetative development. Our finding that several genes are regulated by genomic imprinting in the Arabidopsis embryo clearly demonstrates that this epigenetic phenomenon is not a unique feature of the endosperm in both monocots and dicots.

Regulatory Elements Associated with Paternally-Expressed Genes in the Imprinted Murine Angelman/Prader-Willi Syndrome Domain

The Angelman/Prader-Willi syndrome (AS/PWS) domain contains at least 8 imprinted genes regulated by a bipartite imprinting center (IC) associated with the SNRPN gene. One component of the IC, the PWS-IC, governs the paternal epigenotype and expression of paternal genes. The mechanisms by which imprinting and expression of paternal genes within the AS/PWS domain – such as MKRN3 and NDN – are regulated by the PWS-IC are unclear. The syntenic region in the mouse is organized and imprinted similarly to the human domain with the murine PWS-IC defined by a 6 kb interval within the Snrpn locus that includes the promoter. To identify regulatory elements that may mediate PWS-IC function, we mapped the location and allele-specificity of DNase I hypersensitive (DH) sites within the PWS-IC in brain cells, then identified transcription factor binding sites within a subset of these DH sites. Six major paternal-specific DH sites were detected in the Snrpn gene, five of which map within the 6 kb PWS-IC. We postulate these five DH sites represent functional components of the murine PWS-IC. Analysis of transcription factor binding within multiple DH sites detected nuclear respiratory factors (NRF's) and YY1 specifically on the paternal allele. NRF's and YY1 were also detected in the paternal promoter region of the murine Mrkn3 and Ndn genes. These results suggest that NRF's and YY1 may facilitate PWS-IC function and coordinately regulate expression of paternal genes. The presence of NRF's also suggests a link between transcriptional regulation within the AS/PWS domain and regulation of respiration. 3C analyses indicated Mkrn3 lies in close proximity to the PWS-IC on the paternal chromosome, evidence that the PWS-IC functions by allele-specific interaction with its distal target genes. This could occur by allele-specific co-localization of the PWS-IC and its target genes to transcription factories containing NRF's and YY1.

Mechanisms of activation of the paternally expressed genes by the Prader-Willi imprinting center in the Prader-Willi/Angelman syndromes domains

The Prader-Willi syndrome/Angelman syndrome (PWS/AS) imprinted domain is regulated by a bipartite imprinting control center (IC) composed of a sequence around the SNRPN promoter (PWS-IC) and a 880-bp sequence located 35 kb upstream (AS-IC). The AS-IC imprint is established during gametogenesis and confers repression upon PWS-IC on the maternal allele. Mutation at PWS-IC on the paternal allele leads to gene silencing across the entire PWS/AS domain. This silencing implies that PWS-IC functions on the paternal allele as a bidirectional activator. Here we examine the mechanism by which PWS-IC activates the paternally expressed genes (PEGs) using transgenes that include the PWS-IC sequence in the presence or absence of AS-IC and NDN, an upstream PEG, as an experimental model. We demonstrate that PWS-IC is in fact an activator of NDN. This activation requires an unmethylated PWS-IC in the gametes and during early embryogenesis. PWS-IC is dispensable later in development. Interestingly, a similar activation of a nonimprinted gene (APOA1) was observed, implying that PWS-IC is a universal activator. To decipher the mechanism by which PWS-IC confers activation of remote genes, we performed methylated DNA immunoprecipitation (MeDIP) array analysis on lymphoblast cell lines that revealed dispersed, rather than continued differential methylation. However, chromatin conformation capture (3c) experiments revealed a physical interaction between PWS-IC and the PEGs, suggesting that activation of PEGs may require their proximity to PWS-IC.

Lacking expression of paternally-expressed gene confirms the failure of syngamy after intracytoplasmic sperm injection in swamp buffalo (Bubalus bubalis)

The objectives were to: (1) examine the efficiency of intracytoplasmic sperm injection (ICSI) technique, with or without chemical activation of in vitro matured buffalo oocytes, on sperm head decondensation; and (2) compare the subsequent development of embryos after activation of ICSI (ICSI (+) activation group) and sham injection (Sham (+) activation group) oocytes (embryos obtained by in vitro fertilization of IVM oocytes served as a control group). Pronuclear formation rates in ICSI (+) activation and Sham (+) activation groups were higher than that of ICSI without activation (P < 0.05). However, because 90.9% of presumptive zygotes in ICSI (+) activation group demonstrated pronuclear formation with an intact sperm head, we inferred that most were parthenotes. Neither developmental competence (morula and blastocyst formation rates) nor mean total cell number of blastocysts was significantly different among ICSI (+) activation, Sham (+) activation, and IVF groups. To clarify whether blastocysts were derived from syngamy or parthenogenesis, expression of Nnat, a paternally expressed gene in blastocysts derived from IVF, ICSI and oocyte activation without sperm or sham injection was additionally examined using reverse transcription polymerase chain reaction (RT-PCR). Expression of Nnat mRNA was not detected in ICSI (+) activation blastocysts, indicating failure of male genome activation. Although blastocyst development after ICSI combined with chemical activation was similar to IVF oocytes, these blastocysts were generated by parthenogenesis, due to failure of male pronucleus formation.

Parent-of-Origin Effects on Gene Expression and DNA Methylation in the Maize Endosperm

Imprinting describes the differential expression of alleles based on their parent of origin. Deep sequencing of RNAs from maize (Zea mays) endosperm and embryo tissue 14 d after pollination was used to identify imprinted genes among a set of ~12,000 genes that were expressed and contained sequence polymorphisms between the B73 and Mo17 genotypes. The analysis of parent-of-origin patterns of expression resulted in the identification of 100 putative imprinted genes in maize endosperm, including 54 maternally expressed genes (MEGs) and 46 paternally expressed genes (PEGs). Three of these genes have been previously identified as imprinted, while the remaining 97 genes represent novel imprinted maize genes. A genome-wide analysis of DNA methylation identified regions with reduced endosperm DNA methylation in, or near, 19 of the 100 imprinted genes. The reduced levels of DNA methylation in endosperm are caused by hypomethylation of the maternal allele for both MEGs and PEGs in all cases tested. Many of the imprinted genes with reduced DNA methylation levels also show endosperm-specific expression patterns. The imprinted maize genes were compared with imprinted genes identified in genome-wide screens of rice (Oryza sativa) and Arabidopsis thaliana, and at least 10 examples of conserved imprinting between maize and each of the other species were identified.

High-resolution analysis of parent-of-origin allelic expression in the Arabidopsis Endosperm.

Genomic imprinting is an epigenetic phenomenon leading to parent-of-origin specific differential expression of maternally and paternally inherited alleles. In plants, genomic imprinting has mainly been observed in the endosperm, an ephemeral triploid tissue derived after fertilization of the diploid central cell with a haploid sperm cell. In an effort to identify novel imprinted genes in Arabidopsis thaliana, we generated deep sequencing RNA profiles of F1 hybrid seeds derived after reciprocal crosses of Arabidopsis Col-0 and Bur-0 accessions. Using polymorphic sites to quantify allele-specific expression levels, we could identify more than 60 genes with potential parent-of-origin specific expression. By analyzing the distribution of DNA methylation and epigenetic marks established by Polycomb group (PcG) proteins using publicly available datasets, we suggest that for maternally expressed genes (MEGs) repression of the paternally inherited alleles largely depends on DNA methylation or PcG-mediated repression, whereas repression of the maternal alleles of paternally expressed genes (PEGs) predominantly depends on PcG proteins. While maternal alleles of MEGs are also targeted by PcG proteins, such targeting does not cause complete repression. Candidate MEGs and PEGs are enriched for cis-proximal transposons, suggesting that transposons might be a driving force for the evolution of imprinted genes in Arabidopsis. In addition, we find that MEGs and PEGs are significantly faster evolving when compared to other genes in the genome. In contrast to the predominant location of mammalian imprinted genes in clusters, cluster formation was only detected for few MEGs and PEGs, suggesting that clustering is not a major requirement for imprinted gene regulation in Arabidopsis.

An intronic DNA sequence within the mouse Neuronatin gene exhibits biochemical characteristics of an ICR and acts as a transcriptional activator in Drosophila

Imprinting control regions (ICRs) are domains within imprinted loci that are essential for their establishment and maintenance. Imprinted loci can extend over several megabases, encompass both maternally and paternally-expressed genes and exhibit multiple and complex epigenetic modifications including large regions of allele-specific DNA methylation. Differential chromatin organisation has also been observed within imprinted loci but is restricted to the ICRs. In this study we report the identification of a novel imprinting control region for the mouse Neuronatin gene. This biochemically defined putative ICR, present within its 250 bp second intron, functions as transcriptional activator in Drosophila. This is unlike other known ICRs which have been shown to function as transcriptional silencers. Furthermore, at the endogenous locus, the activating signal from the ICR extends to the Neuronatin promoter via allele-specific unidirectional nucleosomal positioning. Our results support the proposal that the Neuronatin locus employs the most basic mechanism for establishing allele-specific gene expression and could provide the foundation for the multiplex arrangements reported at more complex loci.

Deletions and epimutations affecting the human 14q32.2 imprinted region in individuals with paternal and maternal upd(14)-like phenotypes

Human chromosome 14q32.2 carries a cluster of imprinted genes including paternally expressed genes (PEGs) such as DLK1 and RTL1 and maternally expressed genes (MEGs) such as MEG3 (also known as GTL2), RTL1as (RTL1 antisense) and MEG8 (refs. 1,2), together with the intergenic differentially methylated region (IG-DMR) and the MEG3-DMR. Consistent with this, paternal and maternal uniparental disomy for chromosome 14 (upd(14)pat and upd(14)mat) cause distinct phenotypes. We studied eight individuals (cases 1-8) with a upd(14)pat-like phenotype and three individuals (cases 9-11) with a upd(14)mat-like phenotype in the absence of upd(14) and identified various deletions and epimutations affecting the imprinted region. The results, together with recent mouse data, imply that the IG-DMR has an important cis-acting regulatory function on the maternally inherited chromosome and that excessive RTL1 expression and decreased DLK1 and RTL1 expression are relevant to upd(14)pat-like and upd(14)mat-like phenotypes, respectively.

Molecular mechanisms leading to the phenotypic development in paternal and maternal uniparental disomy for chromosome 14.

Human chromosome 14q32.2 carries a cluster of imprinted genes. They include paternally expressed genes (PEGs) such as DLK1 and RTL1, and maternally expressed genes (MEGs) such as GTL2 (alias, MEG3), RTL1as (RTL1 antisense), and MEG8. Consistent with this, paternal and maternal uniparental disomies for chromosome 14 (upd(14)pat and upd(14)mat) cause distinct phenotypes. In this review, we summarize the current knowledge about the underlying factors for the development of upd(14)pat and upd(14)mat phenotypes. The data available suggest that the DLK1-GTL2 intergenic differentially methylated region (IG-DMR) plays an important role in the maternal to paternal epigenotypic switch, and that excessive RTL1 expression and decreased DLK1 and RTL1 expression play a major role in the development of upd(14)pat-like and upd(14)mat-like phenotypes, respectively.

Neuronally-expressed necdin gene: an imprinted candidate gene in Prader-Willi syndrome

Prader-Willi syndrome (PWS) is associated with paternal-specific deficiencies of human chromosome 15q11–q13, caused by paternal deletion, maternal uniparental disomy (UPD), or imprinting mutations. Several imprinted, paternally-expressed genes have been identified within the ∼1–1·5 Mb PWS candidate region, including SNRPN, IPW, ZNF127, PAR-5, PAR-1, and PAR-SN. 1 Sutcliffe JS Jiang YH Galjaard RJ et al. The E6–AP ubiquitin-protein ligase (UbeSa) gene is localized within a narrowed Angelman syndrome critical region. Genome Res. 1997; 7: 368-377 PubMed Google Scholar Because the three major classes of PWS result in the loss of all paternal-specific gene expression, PWS may well represent a contiguous gene syndrome, with deficiencies of several genes contributing to the phenotype.