Mechanisms Of Epigenetic Reprogramming Research Articles

Abstract 859: Development of non-smoking lung cancers by indoor carcinogenic aerosols through epigenetic reprogramming of lung stem cells: Bioinformatics and artificial intelligence analysis

Abstract Several scientific studies discovered the toxic effects of indoor aerosols on different organs’ adult stem cells of the human body. The most importantly, the indoor toxic aerosols comprised of numerous carcinogens namely, sodium dodecyl sulfate, phthalic acids from floor cleaners, formaldehyde, acetaldehyde, benzene, toluene, ethyl benzene, xylenes, from air fresheners, Hydramethylnon, Pyrethrins, imiprothrin, cypermethrin, tetramethrin, prallethrin and permethrin from ant and roach Killer. However, there was a lack of analysis revealing the mechanism of toxic effects and epigenetic reprogramming of human lung stem cells and development of non-smoking lung cancers. In this study, we analyzed the toxic effects of these chemicals and the mechanism of epigenetic reprograming on lung epithelial cells using Comparative Toxico-genomics Database (CTD), database of Environmental Health Perspectives (EHP), National Institute of Environmental Health Sciences (NIEHS), Google search browser with generative artificial intelligence (AI) and other biological informatics methods (Reactome, Cytoscape PSICQUIC services, and ChEMBL database). We observed from CTD analysis that the chemical gene interactions as risk factors from indoor aerosols showed transcriptionally highly expressed of 31 common genes, by phthalic acids, sodium dodecyl sulfate, from the total of 1347 genes, 107 common genes by formaldehyde, acetaldehyde from the total of 4193 genes and all other toxic ingredients of indoor aerosols. The epigenetic mechanism of histone modifications (H3K4me3, H3K9me3, H3K27me3, H3K27ac, H3K36me3, H3K4me1, Hsa-mir-1301 of lung epithelial, and miRNAs have been revealed by NIH Roadmap Epigenomics Mapping Consortium and the lung tissue arrays were analyzed with AI and matplotlib program. The heterogenic adult stem cell reprogramming is regulated by epigenetic bivalent histone modifications at transcript starts sequences (TSS) of target genes and our current study revealed that CD44, CD80, ALDH2, ALDH3A1, ALDH3B2 are epigenetically reprogrammed with close interaction of other genes by these toxic ingredients of indoor aerosols. Therefore, our results are significant for the carcinogenic effects of indoor aerosols causing for development of lung cancer epigenetically, which may give a clue for the prevention and treatment of nonsmoking lung cancer. Citation Format: Juan Anderson, Mariah Delgado, Malcolm Lovett, Maya Saunders, Geovannie Lake, Samuel Darko, Rose M. Stiffin, Ayivi Huisso, Marilyn Sherman, Latoya Appleton, Amalya Mihnea, Swarnava Das, Mohammad M. Algahtani, Ravi Vadapalli, Biswarup Basu, Arunima Biswas, Noor Neha, Madhumita Das, Marco A. Ruiz, Mayur Doke, Jayanta K. Das. Development of non-smoking lung cancers by indoor carcinogenic aerosols through epigenetic reprogramming of lung stem cells: Bioinformatics and artificial intelligence analysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 859.

An insight into the mechanisms of action of selected bioactive compounds against epigenetic targets of prostate cancer: implications on histones modifications.

Prostate cancer is a leading cause of morbidity and mortality among men globally. In this study, we employed an in silico approach to predict the possible mechanisms of action of selected novel compounds reported against prostate cancer epigenetic targets and their derivatives, exhausting through ADMET profiling, drug-likeness, and molecular docking analyses. The selected compounds: sulforaphane, silibinin, 3, 3'-diindolylmethane (DIM), and genistein largely conformed to ADMET and drug-likeness rules including Lipinski's. Docking studies revealed strong binding energy of sulforaphane with HDAC6 (- 4.2kcal/ mol), DIM versus HDAC2 (- 5.2kcal/mol), genistein versus HDAC6 (- 4.1kcal/mol), and silibinin against HDAC1 (- 7.0kcal/mol) coupled with improved binding affinities and biochemical stabilities after derivatization. Findings from this study may provide insight into the potential epigenetic reprogramming mechanisms of these compounds against prostate cancer and could pave the way toward more success in prostate cancer phytotherapy.

Abstract 6584: Potential carcinogenic effects of electronic nicotine delivery systems through epigenetic reprogramming in oral, brain, and lung stem cells

Abstract Recent scientific studies exhibited that the electronic nicotine delivery systems (ENDS) aerosols comprise numerous carcinogens namely propylene glycol, diacetyl, formaldehyde, acetaldehyde, crotonaldehyde (2-butenal), acetone, acetyl propionyl (2,3-pentanedione), acrolein, nicotine, arsenic, cadmium and radioisotopes, such as Polonium-210 and lead-210. However, there was no such investigation for clarifying the mechanism of carcinogenic effects and epigenetic reprogramming of human oral, brain, and lung stem cells. In this study, we investigated the carcinogenic effects of ENDS aerosols, and the mechanism of epigenetic reprogramming using the Comparative Toxico-genomics Database (CTD), NIH Toxicology Data Network (TOXNET), the database of the National Institute of Environmental Health Sciences (NIEHS), AACR Genomics Evidence Neoplasia Information Exchange (GENIE) database, and other bioinformatics databases. We found from CTD analysis that the chemical gene interactions showed approximately 3% of genes highly expressed by propylene glycol and diacetyl from a total of 98 genes. The AACR GENIE database analysis showed EGFR, is associated with gliomas (191 clinical data, DFCI glioma cohort), found among 98 genes regulated by propylene glycol and diacetyl. The CTD analysis also exhibited the chemical gene interactions of highly expressed 9 common genes (with miR140 by propylene glycol, and formaldehyde from a total of 3983 genes; 5 common genes by propylene glycol, acetaldehyde, from a total of 275 genes; 1 common gene by propylene glycol, crotonaldehyde (2-butenal), from a total of 227 genes; 7 common genes by propylene glycol, acrolein from total 2185 genes; 18 common genes by propylene glycol, and nicotine from total 1244 genes; 11 common genes by propylene glycol, and arsenic from total 4983 genes; 9 common genes with HDAC4, CD86, miR21 by propylene glycol, and cadmium from total 5242 genes. The ENDs aerosols contain Polonium-210 and lead-210 (concentration; 13 ± 2 Bq/kg), which have been reported as Group 1 human carcinogens and emit alpha particles. We found that alpha particles caused DNA lesions, and gene mutations in human oral, brain, and lung stem cells and resulted in long-term chromosomal instability in primary human T lymphocytes by analyzing recent works of literature. The epigenetic mechanisms of histone modifications and these potential carcinogens have been revealed by the database of NIH Roadmap Epigenomics Mapping Consortium and our analysis discovered that Mitogen-activated protein kinase 1(MAPK1) is epigenetically reprogrammed with close interaction of teen genes. Therefore, it is important to find the actual epigenetic mechanism of transcriptional regulation by these potential carcinogens on oral, lung, and brain stem cells, and our current study revealed this important relationship. Citation Format: Akilah Shepard, Oneilia Yearde, Juan Sanchez, Ryan Amador, Gabriel Xirinachs, Erika Hernandez, Elizabeth Corrales, Swarnava Das, Marco Ruiz, Madhumita Das, Rose M. Stiffin, Marilyn Sherman, Alessandra Manzon, Ayivi Huisso, Jayanta K. Das. Potential carcinogenic effects of electronic nicotine delivery systems through epigenetic reprogramming in oral, brain, and lung stem cells. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6584.

Epigenetic regulation of Fructose-1,6-bisphosphatase 1 by host transcription factor Speckled 110 kDa during hepatitis B virus infection.

Hepatitis B virus (HBV) is the leading cause of liver disease ranging from acute and chronic hepatitis to liver cirrhosis and hepatocellular carcinoma (HCC). Studies have revealed that HBV infection broadly reprogrammes the host cellular metabolic processes for viral pathogenesis. Previous reports have shown that glycolysis and gluconeogenesis are among the most deregulated pathways during HBV infection. We noted that despite being one of the rate-limiting enzymes of gluconeogenesis, the role and regulation of Fructose-1,6-bisphosphatase 1 (FBP1) during HBV infection is not much explored. In this study, we report FBP1 upregulation upon HBV infection and unravel a novel mechanism of epigenetic reprogramming of FBP1 by HBV via utilizing host factor Speckled 110 kDa (Sp110). Here, we identified acetylated lysine 18 of histone H3 (H3K18Ac) as a selective interactor of Sp110 Bromodomain. Furthermore, we found that Sp110 gets recruited on H3K18Ac-enriched FBP1 promoter, and facilitates recruitment of deacetylase Sirtuin 2 (SIRT2) on that site in the presence of HBV. SIRT2 in turn brings its interactor and transcriptional activator Hepatocyte nuclear factor 4-alpha to the promoter, which ultimately leads to a loss of DNA methylation near the cognate site. Interestingly, this Sp110 driven FBP1 regulation during infection was found to promote viral-borne HCC progression. Moreover, Sp110 can be used as a prognostic marker for the hepatitis-mediated HCC patients, where high Sp110 expression significantly lowered their survival. Thus, the epigenetic reader protein Sp110 has potential to be a therapeutic target to challenge HBV-induced HCCs.

Abstract 5830: Carcinogenic effects of metal and metalloids through epigenetic reprogramming of human bone marrow stem cells: The bioinformatics database analysis

Abstract Many scientific studies showed that the carcinogenic effects of environmental contamination of several metal and metalloids on bone marrow stem cells of human body. These carcinogens include, Cadmium, Mercury, Chromium, Aluminum, Nickel and Arsenic. However, there was no such analysis for elucidating the mechanism of carcinogenic effects and epigenetic reprogramming of human bone marrow stem cells. In this study, we analyzed the carcinogenic effects of mentioned carcinogens, and the mechanism of epigenetic reprogramming using NIH Toxicology Data Network (TOXNET), Comparative Toxico-genomics Database (CTD), AACR Genomics Evidence Neoplasia Information Exchange (GENIE) database, database of Environmental Health Perspectives (EHP), National Institute of Environmental Health Sciences (NIEHS), and other bioinformatics database. We found from CTD analysis that the chemical gene interactions showed highly expressed of 272 common genes with miR140 by Cadmium, Mercury from total 3704 genes (approximately 7% affected genes). The AACR GENIE database analysis showed CDKN3, is associated with bone cancer (2 clinical data), HDAC7, TP53, MAPK3K1, MAP2K2 (8 clinical data) found among 272 genes regulated by Cadmium and Mercury. The CTD analysis showed the chemical gene interactions of highly expressed of 102 common genes with miR140 by Chromium, Mercury from a total of 1406 genes; 268 common genes with CD84, CD226 by Mercury, Nickel from a total of 5032 genes; 103 common genes with CD226 by Aluminum, Mercury from a total of 2556 genes; 248 common genes by Arsenic, Mercury from total 5508 genes. Among these 5508 genes, 248 common genes with ALDHA1, 247 common genes with CD40, 248 common genes with CD44, 247 common genes with CD423, 247 common genes with miR10A by Arsenic, Mercury from total 2556 genes. The epigenetic mechanisms of histone modifications (H3K9me3, H3K27me3, H3K27ac, H3K36me3, H3K4me1, H3K4me3 of B cells, hemopoietic stem cells, mesenchymal bone marrow, and chondrocyte stem cells and miRNAs) and these carcinogens have been revealed by the database of EHP, NIEHS, the NIH Roadmap Epigenomics Mapping Consortium, the National Center for Biotechnology Information (NCBI), and U.S. National Library of Medicine (NLM) database. Our previous study reported that the regulation of epigenetic bivalent histone modifications at transcript starts sequences (TSS) of target genes for heterogenic adult stem cells (CSCs)reprogramming and our current bioinformatics analysis revealed that TP53 is epigenetically reprogrammed with close interaction of thirteen genes by these carcinogens. Therefore, it is important to find the actual epigenetic mechanism both genomic and epigenomic transcriptional regulation and carcinogenic effects of these carcinogens on bone marrow stem cells, and our current study revealed this important relationship. Citation Format: Benedrica Solomon, Richgena Sturrup, Jada Dixon, Kiyona Jackson, Biancaliz Cruz, Charlissia Ferguson, Taylor Walker, Ralph Elisson, Nathan Harris, Jesennia Bonilla, Rose Mary Stiffin, Marilyn Sherman, Alessandra Manzon, Swarnava Das, Madhumita Das, Jayanta Kumar Das. Carcinogenic effects of metal and metalloids through epigenetic reprogramming of human bone marrow stem cells: The bioinformatics database analysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5830.

Neonatal Exposure to BPA, BDE-99, and PCB Produces Persistent Changes in Hepatic Transcriptome Associated With Gut Dysbiosis in Adult Mouse Livers.

Recent evidence suggests that complex diseases can result from early life exposure to environmental toxicants. Polybrominated diphenyl ethers (PBDEs), and polychlorinated biphenyls (PCBs) are persistent organic pollutants (POPs) and remain a continuing risk to human health despite being banned from production. Developmental BPA exposure mediated-adult onset of liver cancer via epigenetic reprogramming mechanisms has been identified. Here, we investigated whether the gut microbiome and liver can be persistently reprogrammed following neonatal exposure to POPs, and the associations between microbial biomarkers and disease-prone changes in the hepatic transcriptome in adulthood, compared with BPA. C57BL/6 male and female mouse pups were orally administered vehicle, BPA, BDE-99 (a breast milk-enriched PBDE congener), or the Fox River PCB mixture (PCBs), once daily for three consecutive days (postnatal days [PND] 2-4). Tissues were collected at PND5 and PND60. Among the three chemicals investigated, early life exposure to BDE-99 produced the most prominent developmental reprogramming of the gut-liver axis, including hepatic inflammatory and cancer-prone signatures. In adulthood, neonatal BDE-99 exposure resulted in a persistent increase in Akkermansia muciniphila throughout the intestine, accompanied by increased hepatic levels of acetate and succinate, the known products of A. muciniphila. In males, this was positively associated with permissive epigenetic marks H3K4me1 and H3K27, which were enriched in loci near liver cancer-related genes that were dysregulated following neonatal exposure to BDE-99. Our findings provide novel insights that early life exposure to POPs can have a life-long impact on disease risk, which may partly be regulated by the gut microbiome.

Epigenetic reprogramming mechanisms of immunity during influenza A virus infection

This paper reviews epigenetic mechanisms by which influenza viruses affect cellular gene activity to control their life cycles, aiming to provide new insights into the complexity of functional interactions between viral and cellular factors, as well as to introduce novel targets for therapeutic intervention and vaccine development against influenza infections.

The asynchronous establishment of chromatin 3D architecture between in vitro fertilized and uniparental preimplantation pig embryos

BackgroundPigs are important animals for agricultural and biomedical research, and improvement is needed for use of the assisted reproductive technologies. Determining underlying mechanisms of epigenetic reprogramming in the early stage of preimplantation embryos derived from in vitro fertilization (IVF), parthenogenesis, and androgenesis will not only contribute to assisted reproductive technologies of pigs but also will shed light into early human development. However, the reprogramming of three-dimensional architecture of chromatin in this process in pigs is poorly understood.ResultsWe generate three-dimensional chromatin profiles for pig somatic cells, IVF, parthenogenesis, and androgenesis preimplantation embryos. We find that the chromosomes in the pig preimplantation embryos are enriched for superdomains, which are more rare in mice. However, p(s) curves, compartments, and topologically associated domains (TADs) are largely conserved in somatic cells and are gradually established during preimplantation embryogenesis in both mammals. In the uniparental pig embryos, the establishment of chromatin architecture is highly asynchronized at all levels from IVF embryos, and a remarkably strong decompartmentalization is observed during zygotic genome activation (ZGA). Finally, chromosomes originating from oocytes always establish TADs faster than chromosomes originating from sperm, both before and during ZGA.ConclusionsOur data highlight a potential unique 3D chromatin pattern of enriched superdomains in pig preimplantation embryos, an unusual decompartmentalization process during ZGA in the uniparental embryos, and an asynchronized TAD reprogramming between maternal and paternal genomes, implying a severe dysregulation of ZGA in the uniparental embryos in pigs.

Characterization and comparative analyses of transcriptomes of cloned and in vivo fertilized porcine pre-implantation embryos.

ABSTRACTSomatic cell nuclear transfer (SCNT) is the only method known to rapidly reprogram differentiated cells into totipotent embryos. Most cloned embryos become arrested before implantation and the details of the underlying molecular mechanism remain largely unknown. Dynamic regulation of the transcriptome is a key molecular mechanism driving early embryonic development. Here, we report comprehensive transcriptomic analysis of cloned embryos (from Laiwu and Duroc pigs) and in vivo fertilized embryos (from Duroc pigs) using RNA-sequencing. Comparisons between gene expression patterns were performed according to differentially expressed genes, specific-expressed genes, first-expressed genes, pluripotency genes and pathway enrichment analysis. In addition, we closely analyzed the improperly expressed histone lysine methyltransferases and histone lysine demethylases during cell reprogramming in cloned embryos. In summary, we identified altered gene expression profiles in porcine cloned pre-implantation embryos in comparison to normal in vivo embryos. Our findings provide a substantial framework for further discovery of the epigenetic reprogramming mechanisms in porcine SCNT embryos.

Epigenetic Regulation and Its Therapeutic Potential in Pulmonary Hypertension.

Recent advances in epigenetics have made a tremendous impact on our knowledge of biological phenomena and the environmental stressors on complex diseases. Understanding the mechanism of epigenetic reprogramming during the occurrence of pulmonary hypertension (PH) is important for advanced studies and clinical therapy. In this article, we review the discovery of novel epigenetic mechanisms associated with PH including DNA methylation, histone modification, and noncoding RNA interference. In addition, we highlight the role of epigenetic mechanisms in adult PAH resulting from undesirable perinatal environments—Extrauterine growth restriction (EUGR) and Intrauterine growth retardation (IUGR). Lastly, we give a comprehensive summary for the remaining challenges and discuss future methods of epigenetic targeted therapy for pulmonary hypertension.

Use of Histone K-M Mutants for the Analysis of Transcriptional Regulation in Mouse Zygotes.

Histone modifications are dramatically altered during the pronuclear (PN) stage of zygotes, and more markedly in paternal than maternal pronuclei. Among various types of histone modifications, lysine methylation exhibits the most dynamic changes in the PN stage . To analyze the physiological functions of histone methylations, it is therefore important to elucidate the mechanism of epigenetic reprogramming. However, loss-of-function approaches using mutant histones whose lysine residues have been substituted with arginine residues are unable to erase histone modifications at all levels, since they are incapable of entirely replacing endogenous histones. To solve this problem, we used an alternative histone mutant whose lysine residues were substituted with methionine (K-M mutants). This mutant cannot be methylated itself but also prevents methylation of endogenous histones. We also developed a simple method for analyzing global transcription levels in early preimplantation embryos, involving using a commercial kit to examine the involvement of histone methylation in zygotic gene activation.

Zebrafish Discoveries in Cancer Epigenetics.

The cancer epigenome is fundamentally different than that of normal cells. How these differences arise in and contribute to carcinogenesis is not known, and studies using model organisms such as zebrafish provide an opportunity to address these important questions. Modifications of histones and DNA comprise the complex epigenome, and these influence chromatin structure, genome stability and gene expression, all of which are fundamental to the cellular changes that cause cancer. The cancer genome atlas covers the wide spectrum of genetic changes associated with nearly every cancer type, however, this catalog is currently uni-dimensional. As the pattern of epigenetic marks and chromatin structure in cancer cells is described and overlaid on the mutational landscape, the map of the cancer genome becomes multi-dimensional and highly complex. Two major questions remain in the field: (1) how the epigenome becomes repatterned in cancer and (2) which of these changes are cancer-causing. Zebrafish provide a tractable in vivo system to monitor the epigenome during transformation and to identify epigenetic drivers of cancer. In this chapter, we review principles of cancer epigenetics and discuss recent work using zebrafish whereby epigenetic modifiers were established as cancer driver genes, thus providing novel insights into the mechanisms of epigenetic reprogramming in cancer.

P53‐Mediated Repression of the Reprogramming in Cloned Bovine Embryos Through Direct Interaction with HDAC1 and Indirect Interaction with DNMT3A

P53 is a transcriptional activator, regulating growth arrest, DNA repair and apoptosis. We found that the expression level of P53 and the epigenetic profiles were significantly different in bovine somatic cell nuclear transfer embryos from those in vitro fertilization (IVF) embryos. So we inferred that abnormally expression of P53 might contribute to the incomplete reprogramming. Using bovine foetal fibroblasts, we constructed and screened a highly efficient shRNA vector targeting bovine P53 gene and then reconstituted somatic cell nuclear transfer embryos (RNAi-SCNT). The results indicated that expression levels of P53 were downregulated significantly in RNAi-SCNT embryos, and the blastulation rate and the total number of cell increased significantly. Moreover, methylation levels of CpG islands located 5' region of OCT4, NANOG, H19 and IGF2R in RNAi -SCNT embryos were significantly normalized to that IVF embryos, and the methylation levels of genome DNA, H3K9 and H4K5 acetylation levels were also returned to levels similar to the IVF embryos. Differentially expressed genes were identified by microarray, and 28 transcripts were found to be significantly different (> twofolds) in RNAi-SCNT embryos compared to the control nuclear transfer embryos (SCNT). Among the 28 differentially expressed transcripts, just HDAC1 and DNMT3A were closely associated with the epigenetic modifications. Finally, ChIP further showed that P53 might repress the epigenetic reprogramming by regulating HDAC1 directly and DNMT3A indirectly. These findings offer significant references to further elucidate the mechanism of epigenetic reprogramming in SCNT embryos.

Allele-specific DNA methylation analyses associated with siRNAs in Arabidopsis hybrids

Accumulating evidence has suggested that epigenetic marks including DNA methylation, small RNA and histone modification may involve hybrid vigor in plants. However, knowledge about how epigenetic marks in hybrids regulate gene expression is still limited. Based on genome-wide DNA methylation landscapes of Arabidopsis thaliana Ler and C24 ecotypes and their reciprocal F1 hybrids which were obtained in our previous work, we analyzed allele-specific DNA methylation and distinguished cis- and trans-regulated DNA methylation in hybrids. Our study indicated that both cis- and trans-regulated DNA methylation played roles in hybrids, when cis-regulation played a major role in CG methylation and trans-regulation played major roles in CHG and CHH methylation. In addition, we observed correlations between trans-regulated DNA methylation and siRNA densities. Enriched siRNA regions were significantly concurrent with highly trans-regulated DNA methylation regions. Our results illustrated DNA methylation regulation patterns integrated with siRNAs in Arabidopsis hybrids, and shed light on understanding the mechanism of epigenetic reprogramming for hybrid vigor.

EPIGENETIC VARIATION IN ASEXUALLY REPRODUCING ORGANISMS

The role that epigenetic inheritance can play in adaptation may differ between sexuals and asexuals because (1) the dynamics of adaptation differ under sexual and asexual reproduction and the opportunities offered by epigenetic inheritance may affect these dynamics differently; and (2) in asexual reproduction epigenetic reprogramming mechanisms that are associated with meiosis can be bypassed, which could promote the buildup of epigenetic variation in asexuals. Here, we evaluate current evidence for an epigenetic contribution to adaptation in asexuals. We argue that two aspects of epigenetic variation should have particular relevance for asexuals, namely epigenetics-mediated phenotypic plasticity within and between generations, and heritable variation via stochastic epimutations. An evaluation of epigenetic reprogramming mechanisms suggests that some, but not all, forms of asexual reproduction enhance the likelihood of stable transmission of epigenetic marks across generations compared to sexual reproduction. However, direct tests of these predicted sexual-asexual differences are virtually lacking. Stable transmission of DNA methylation, transcriptomes, and phenotypes from parent to clonal offspring are demonstrated in various asexual species, and clonal genotypes from natural populations show habitat-specific DNA methylation. We discuss how these initial observations can be extended to demonstrate an epigenetic contribution to adaptation.

Postnatal epigenetic reprogramming in the germline of a marsupial, the tammar wallaby

BackgroundEpigenetic reprogramming is essential to restore totipotency and to reset genomic imprints during mammalian germ cell development and gamete formation. The dynamic DNA methylation change at DMRs (differentially methylated regions) within imprinted domains and of retrotransposons is characteristic of this process. Both marsupials and eutherian mammals have genomic imprinting but these two subgroups have been evolving separately for up to 160 million years. Marsupials have a unique reproductive strategy and deliver tiny, altricial young that complete their development within their mother's pouch. Germ cell proliferation in the genital ridge continues after birth in the tammar wallaby (Macropus eugenii), and it is only after 25 days postpartum that female germ cells begin to enter meiosis and male germ cells begin to enter mitotic arrest. At least two marsupial imprinted loci (PEG10 and H19) also have DMRs. To investigate the evolution of epigenetic reprogramming in the marsupial germline, here we collected germ cells from male pouch young of the tammar wallaby and analysed the methylation status of PEG10 and H19 DMR, an LTR (long terminal repeat) and a non-LTR retrotransposons.ResultsDemethylation of the H19 DMR was almost completed by 14 days postpartum and de-novo methylation started from 34 days postpartum. These stages correspond to 14 days after the completion of primordial germ cell migration into genital ridge (demethylation) and 9 days after the first detection of mitotic arrest (re-methylation) in the male germ cells. Interestingly, the PEG10 DMR was already unmethylated at 7 days postpartum, suggesting that the timing of epigenetic reprogramming is not the same at all genomic loci. Retrotransposon methylation was not completely removed after the demethylation event in the germ cells, similar to the situation in the mouse.ConclusionsThus, despite the postnatal occurrence of epigenetic reprogramming and the persistence of genome-wide undermethylation for 20 days in the postnatal tammar, the relative timing and mechanism of germ cell reprogramming are conserved between marsupials and eutherians. We suggest that the basic mechanism of epigenetic reprogramming had already been established before the marsupial-eutherian split and has been faithfully maintained for at least 160 million years and may reflect the timing of the onset of mitotic arrest in the male germline.

Effects of DNMT1 and HDAC Inhibitors on Gene-Specific Methylation Reprogramming during Porcine Somatic Cell Nuclear Transfer

Somatic cell nuclear transfer (SCNT) in mammalian cloning currently remains inefficient. Incomplete or erroneous epigenetic reprogramming of specialized donor somatic nuclear and resulting aberrant gene expression during development of cloned embryos is commonly believed as the main reason that causes the low efficiency of SCNT. Use of small molecular reprogramming modifiers to assist the somatic nucleus to mimic naturally occurring DNA methylation and chromatin remodeling in nucleus of fertilization-derived zygotes, has been widely attempted to improve cloning efficiency. However, impacts of these small modifiers on gene-specific methylation dynamics and their potential effects on methylation of imprinted gene have rarely been traced. Here, we attempted two relatively novel DNMT1 inhibitor (DNMTi) and histone deacetylase inhibitor (HDACi), scriptaid and RG108, and demonstrated their effects on dynamics of gene-specific DNA methylation and transcription of porcine SCNT embryos. We found that scriptaid and RG108 had synergetic effects on rescuing the disrupted methylation imprint of H19 during SCNT at least partially by repression over-expressed MBD3 in eight-cell cloned embryos. Furthermore, we firstly identified a differential methylation regions (DMRs) at 5′ flanking regions of XIST gene and found that scriptaid alone and its combination with RG108 modify the dynamics of both transcription and DNA methylation levels in cloned embryos, by different manners. Additionally, we found that scriptaid alone and its combination with RG108 can significantly promote the transcription of NANOG in cloned embryos and enhance their pre-implantation developmental capacity. Our results would contribute to uncovering the epigenetic reprogramming mechanisms underlying the effects of assisted small molecules on improvement of mammalian cloning efficiency.

The simplest explanation: passive DNA demethylation in PGCs.

Primordial germ cell (PGC) specification is one of two major developmental windows where modifications associated with global heterochromatin maintenance are erased. One of the most intriguing and confounding dynamics to rectify has been the apparent global depletion of cytosine methylation, which has been intensely scrutinized for nearly two decades. While numerous reports have suggested active catalytic removal as the primary mechanism, work by Saitou and colleagues presented in this issue of The EMBO Journal provide support for a simpler model whereby the downregulation of essential recruitment factors appears sufficient to erase this mark passively during a phase of rapid proliferation. During mammalian development, transitions in cellular programs are predominantly accompanied by focal changes to chromatin that reflects local activities of transcriptional activators or repressors. As such, the fraction of the genome that is targeted for remodelling during any given transcription factor driven programming event is comparatively small (Dunham et al , 2012). Two exceptions to this general paradigm occur upon specification of the PGC lineage within the developing embryo and upon fertilization within the protamine‐compacted paternal genome (Saitou et al , 2012). In these contexts, dramatic global epigenetic modification appears to restructure the majority of the genome and does so in a rapid and coordinated fashion (Seki et al , 2005). As part of either process, the mechanism behind the apparent global erasure of DNA methylation has posed a particularly vexing problem. In PGCs, this global demethylation is considered essential to erase somatic methylation signatures as well as parental imprints while simultaneously reactivating transcriptional programs associated with pluripotency and gametogenesis. Until recently, the rapidity of DNA demethylation during PGC specification appeared to support numerous mechanisms that largely centred on an active catalytic step, including strategies employing deamination or oxidation followed by base excision repair (Wu and Zhang, 2010). While these proposed mechanisms appeared …

Epigenetic Re-Programming during Mammalian Preimplantation Embryogenesis and PGC Development

Epigenetic programming and reprogramming, by means of DNA methylation and histone modifications etc., control the mammalian development to a large extent. They are also artificially altered for cell fate conversion and regeneration. Though epigenetic modifications change with slow dynamics during somatic cell lineage differentiation, they undergo a genome-wide dramatic change with extensive DNA de-methylation and histone modification during two specific time windows, the early embryogenesis and the Primordial Germ Cell (PGC) development stage. Here we reviewed these global epigenetic reprogramming occurred during normal development, mainly focusing on DNA methylation, histone modification and X-chromosome inactivation. Epigenetic reprogramming participates in many key biological processes such as genomic imprinting, X-chromosome inactivation, gene expression regulation and tumorigenesis and genome stability. Understanding the mechanisms of epigenetic reprogramming during the early embryogenesis and PGC formation would facilitate our knowledge of the developmental process and disease progress.

Epigenetic reprogramming in mouse pre-implantation development and primordial germ cells

Epigenetic modifications are crucial for the identity and stability of cells, and, when aberrant, can lead to disease. During mouse development, the genome-wide epigenetic states of pre-implantation embryos and primordial germ cells (PGCs) undergo extensive reprogramming. An improved understanding of the epigenetic reprogramming mechanisms that occur in these cells should provide important new information about the regulation of the epigenetic state of a cell and the mechanisms of induced pluripotency. Here, we discuss recent findings about the potential mechanisms of epigenetic reprogramming, particularly genome-wide DNA demethylation, in pre-implantation mouse embryos and PGCs.