scholarly journals Alteration of the DNA methylation status of donor cells impairs the developmental competence of porcine cloned embryos

2016 ◽  
Vol 62 (1) ◽  
pp. 71-77 ◽  
Author(s):  
Yan Jun HUAN ◽  
Zhan Feng WU ◽  
Ji Guang ZHANG ◽  
Jiang ZHU ◽  
Bing Teng XIE ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Methylation Status ◽  
Developmental Competence ◽  
Donor Cells

scholarly journals DNA methylation and mRNA expression profiles in bovine oocytes derived from prepubertal and adult donors

Reproduction ◽  
2012 ◽  
Vol 144 (3) ◽  
pp. 319-330 ◽  
Author(s):  
Mike Diederich ◽  
Tamara Hansmann ◽  
Julia Heinzmann ◽  
Brigitte Barg-Kues ◽  
Doris Herrmann ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Mrna Expression ◽  
Bos Taurus ◽  
Expression Profiles ◽  
Methylation Status ◽  
Transcript Abundance ◽  
Developmental Competence ◽  
Satellite Sequences ◽  
Bovine Oocytes

The developmental capacity of oocytes from prepubertal cattle is reduced compared with their adult counterparts, and epigenetic mechanisms are thought to be involved herein. Here, we analyzed DNA methylation in three developmentally important, nonimprinted genes (SLC2A1, PRDX1, ZAR1) and two satellite sequences, i.e. ‘bovine testis satellite I’ (BTS) and ‘Bos taurus alpha satellite I’ (BTαS). In parallel, mRNA expression of the genes was determined by quantitative real-time PCR. Oocytes were retrieved from prepubertal calves and adult cows twice per week over a 3-week period by ultrasound-guided follicular aspiration after treatment with FSH and/or IGF1. Both immature and in vitro matured prepubertal and adult oocytes showed a distinct hypomethylation profile of the three genes without differences between the two types of donors. The methylation status of the BTS sequence changed according to the age and treatment while the methylation status of BTαS sequence remained largely unchanged across the different age and treatment groups. Relative transcript abundance of the selected genes was significantly different in immature and in vitro matured oocytes; only minor changes related to origin and treatment were observed. In conclusion, methylation levels of the investigated satellite sequences were high (>50%) in all groups and showed significant variation depending on the age, treatment, or in vitro maturation. To what extent this is involved in the acquisition of developmental competence of bovine oocytes needs further study.

scholarly journals Epigenetic alteration of the donor cells does not recapitulate the reprogramming of DNA methylation in cloned embryos

Reproduction ◽  
2007 ◽  
Vol 134 (6) ◽  
pp. 781-787 ◽  
Author(s):  
Gabbine Wee ◽  
Jung-Jae Shim ◽  
Deog-Bon Koo ◽  
Jung-Il Chae ◽  
Kyung-Kwang Lee ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Trichostatin A ◽  
Dna Methyltransferases ◽  
Epigenetic Reprogramming ◽  
Preimplantation Development ◽  
Developmental Competence ◽  
Epigenetic Mark ◽  
Satellite Sequence ◽  
Donor Cells

Epigenetic reprogramming is a prerequisite process during mammalian development that is aberrant in cloned embryos. However, mechanisms that evolve abnormal epigenetic reprogramming during preimplantation development are unclear. To trace the molecular event of an epigenetic mark such as DNA methylation, bovine fibroblasts were epigeneticallyaltered by treatment with trichostatin A (TSA) and then individually transferred into enucleated bovine oocytes. In the TSA-treated cells, expression levels of histone deacetylases and DNA methyltransferases were reduced, but the expression level of histone acetyltransferases such as Tip60 and histone acetyltransferase 1 (HAT1) did not change compared with normal cells. DNA methylation levels of non-treated (normal) and TSA-treated cells were 64.0 and 48.9% in the satellite I sequence (P < 0.05) respectively, and 71.6 and 61.9% in the α-satellite sequence respectively. DNA methylation levels of nuclear transfer (NT) and TSA-NT blastocysts in the satellite I sequence were 67.2 and 42.2% (P < 0.05) respectively, which was approximately similar to those of normal and TSA-treated cells. In the α-satellite sequence, NT and TSA-NT embryos were substantially demethylated at the blastocyst stage as IVF-derived embryos were demethylated. The in vitro developmental rate (46.6%) of TSA-NT embryos that were individually transferred with TSA-treated cells was higher than that (31.7%) of NT embryos with non-treated cells (P < 0.05). Our findings suggest that the chromatin of a donor cell is unyielding to the reprogramming of DNA methylation during preimplantation development, and that alteration of the epigenetic state of donor cells may improve in vitro developmental competence of cloned embryos.

38 EFFECTS OF 5-AZACYTIDINE ON DNA HYPERMETHYLATION STATUS OF CLONED MINIATURE PIG EMBRYO

2009 ◽  
Vol 21 (1) ◽  
pp. 119
Author(s):  
H. S. Kim ◽  
Y. I. Jeong ◽  
J. H. Kim ◽  
J. Choi ◽  
Y. W. Jeong ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Methylation Level ◽  
Methylation Status ◽  
Cell Mass ◽  
Somatic Cells ◽  
Abnormal Development ◽  
Control Group ◽  
Blastocyst Development ◽  
Cell Stage ◽  
Donor Cells

During somatic cell nuclear transfer, somatic cells should undergo epigenetic reprogramming in order to attain successful totipotency. Although there are many reasons of low efficiency of cloning, the aberrant DNA methylation status in donor cells is thought to reduce efficiency and increase abnormalities in cloned embryos. The methylation level of cloned embryos is higher than that of in vivo-produced or in vitro-fertilized embryos before occurring de novo methylation. This hyper DNA methylation status has been considered as a reason for the abnormal development of cloned embryos and the related decrease in term number. The aim of this research is to validate the DNA methylation pattern in cloned porcine embryos and to confirm whether reduction of hyper DNA methylation levels results in an improvement in cloning efficiency. First, immunostaining was used to evaluate the stage-specific changing pattern of DNA methylation of cloned embryos. In this result, we demonstrated that the 1-cell stage embryos and blastocyst had the highest level of methylation compared to 2- or 4-cell stage embryos. Second, this methylation level was higher in SCNT-derived embryos compared to parthenogenic embryos, suggesting that the methylation level was associated with the nucleus of donor cells. Third, cloned embryos were treated with various concentration of 5-azacytidine, which is a demethylating agent, to find adequate concentration. In results, 500 nm of 5-azacytidin group from 0 to 24 h after activation produced significantly more blastocysts compared to control group (control; 5.4 ± 2.3, 500 nm/0 24-h treatment; 12.7 ± 2.1, P < 0.05) and also showed low DNA methylation level of inner cell mass different from that of control group. In conclusion, our results suggest that cloned porcine embryos show typical demethylation. But, they are generally on a hyper level of DNA methylation. This high DNA methylation level is associated with somatic cells and affects on blastocyst development. We also suggest that 5-azacytidine could improve blastocyst development rate of cloned porcine embryos by aiding weak and abnormal demethylation process.

Gene-specific profiling of DNA methylation and mRNA expression in bovine oocytes derived from follicles of different size categories

2017 ◽  
Vol 29 (10) ◽  
pp. 2040 ◽  
Author(s):  
F. Mattern ◽  
J. Heinzmann ◽  
D. Herrmann ◽  
A. Lucas-Hahn ◽  
T. Haaf ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Mrna Expression ◽  
Candidate Genes ◽  
Growth Phase ◽  
Methylation Status ◽  
Developmental Competence ◽  
Follicular Growth ◽  
Antral Follicles ◽  
Follicle Diameter ◽  
Bovine Oocytes

Epigenetic changes, such as DNA methylation, play an essential role in the acquisition of full developmental competence by mammalian oocytes during the late follicular growth phase. Here we used the bovine model to investigate the DNA methylation profiles of seven candidate genes (imprinted: bH19, bSNRPN; non-imprinted: bZAR1, bDNMT3A, bOCT4, bDNMT3 Lo and bDNMT3 Ls) and the mRNA expression of nine candidate genes (imprinted: bSNRPN, bPEG3, bIGF2R; non-imprinted: bPRDX1, bDNMT1B, bDNMT3A, bZAR1, bHSF1 and bNLRP9) in oocytes from antral follicles of three different size classes (≤2 mm, 3–5 mm, ≥6 mm) to unravel the epigenetic contribution to this process. We observed an increased number of aberrantly methylated alleles in bH19, bSNRPN and bDNMT3 Lo of oocytes from small antral follicles (≤2 mm), correlating with lower developmental competence. Furthermore, we detected an increased frequency of CpG sites with an unclear methylation status for DNMT3 Ls, specifically in oocytes from follicles ≥6 mm, predominantly at three CpG positions (CpG2, CpG7 and CpG8), of which CpG7 is a potential regulatory site. No major differences in mRNA expression were observed, indicating that the transcriptional machinery may not yet be active during the follicular growth phase. Our results support the notion that a follicle diameter of ~2 mm is a critical stage for establishing DNA methylation profiles and indicate a link between DNA methylation and the acquisition of oocyte developmental competence.

DNA Methylation Status of Mash2 in Lungs of Somatic Cell Cloning Bovines*

2010 ◽  
Vol 37 (9) ◽  
pp. 960-966 ◽  
Author(s):  
Jie CHEN ◽  
Dong-Jie LI ◽  
Cui ZHANG ◽  
Ning LI ◽  
Shi-Jie LI
Keyword(s):  
Dna Methylation ◽  
Somatic Cell ◽  
Methylation Status ◽  
Cell Cloning

scholarly journals Epigenetic Changes During Mechanically Induced Osteogenic Lineage Commitment

2015 ◽  
Vol 137 (2) ◽  
Author(s):  
Julia C. Chen ◽  
Mardonn Chua ◽  
Raymond B. Bellon ◽  
Christopher R. Jacobs
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Shear Stress ◽  
Fluid Shear Stress ◽  
Methylation Status ◽  
Lineage Commitment ◽  
Fluid Shear ◽  
Osteogenic Lineage ◽  
Osteogenic Markers ◽  
Heterochromatin Structure

Osteogenic lineage commitment is often evaluated by analyzing gene expression. However, many genes are transiently expressed during differentiation. The availability of genes for expression is influenced by epigenetic state, which affects the heterochromatin structure. DNA methylation, a form of epigenetic regulation, is stable and heritable. Therefore, analyzing methylation status may be less temporally dependent and more informative for evaluating lineage commitment. Here we analyzed the effect of mechanical stimulation on osteogenic differentiation by applying fluid shear stress for 24 hr to osteocytes and then applying the osteocyte-conditioned medium (CM) to progenitor cells. We analyzed gene expression and changes in DNA methylation after 24 hr of exposure to the CM using quantitative real-time polymerase chain reaction and bisulfite sequencing. With fluid shear stress stimulation, methylation decreased for both adipogenic and osteogenic markers, which typically increases availability of genes for expression. After only 24 hr of exposure to CM, we also observed increases in expression of later osteogenic markers that are typically observed to increase after seven days or more with biochemical induction. However, we observed a decrease or no change in early osteogenic markers and decreases in adipogenic gene expression. Treatment of a demethylating agent produced an increase in all genes. The results indicate that fluid shear stress stimulation rapidly promotes the availability of genes for expression, but also specifically increases gene expression of later osteogenic markers.

scholarly journals DNA methylation differs extensively between strains of the same geographical origin and changes with age in Daphnia magna

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Jack Hearn ◽  
Fiona Plenderleith ◽  
Tom J. Little
Keyword(s):  
Dna Methylation ◽  
Caloric Restriction ◽  
Daphnia Magna ◽  
Single Gene ◽  
Methylation Status ◽  
Strain Differences ◽  
Life History Trait ◽  
Food Level ◽  
Epigenetic Clock ◽  
Age Related

Abstract Background Patterns of methylation influence lifespan, but methylation and lifespan may also depend on diet, or differ between genotypes. Prior to this study, interactions between diet and genotype have not been explored together to determine their influence on methylation. The invertebrate Daphnia magna is an excellent choice for testing the epigenetic response to the environment: parthenogenetic offspring are identical to their siblings (making for powerful genetic comparisons), they are relatively short lived and have well-characterised inter-strain life-history trait differences. We performed a survival analysis in response to caloric restriction and then undertook a 47-replicate experiment testing the DNA methylation response to ageing and caloric restriction of two strains of D. magna. Results Methylated cytosines (CpGs) were most prevalent in exons two to five of gene bodies. One strain exhibited a significantly increased lifespan in response to caloric restriction, but there was no effect of food-level CpG methylation status. Inter-strain differences dominated the methylation experiment with over 15,000 differently methylated CpGs. One gene, Me31b, was hypermethylated extensively in one strain and is a key regulator of embryonic expression. Sixty-one CpGs were differentially methylated between young and old individuals, including multiple CpGs within the histone H3 gene, which were hypermethylated in old individuals. Across all age-related CpGs, we identified a set that are highly correlated with chronological age. Conclusions Methylated cytosines are concentrated in early exons of gene sequences indicative of a directed, non-random, process despite the low overall DNA methylation percentage in this species. We identify no effect of caloric restriction on DNA methylation, contrary to our previous results, and established impacts of caloric restriction on phenotype and gene expression. We propose our approach here is more robust in invertebrates given genome-wide CpG distributions. For both strain and ageing, a single gene emerges as differentially methylated that for each factor could have widespread phenotypic effects. Our data showed the potential for an epigenetic clock at a subset of age positions, which is exciting but requires confirmation.

scholarly journals DNA methylome signatures of prenatal exposure to synthetic glucocorticoids in hippocampus and peripheral whole blood of female guinea pigs in early life

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Aya Sasaki ◽  
Margaret E. Eng ◽  
Abigail H. Lee ◽  
Alisa Kostaki ◽  
Stephen G. Matthews
Keyword(s):  
Dna Methylation ◽  
Whole Blood ◽  
Guinea Pigs ◽  
Critical Role ◽  
Methylation Status ◽  
Peripheral Tissues ◽  
Epigenetic Biomarkers ◽  
Differentially Methylated Genes ◽  
Increased Risk ◽  
Synthetic Glucocorticoids

AbstractSynthetic glucocorticoids (sGC) are administered to women at risk of preterm delivery, approximately 10% of all pregnancies. In animal models, offspring exposed to elevated glucocorticoids, either by administration of sGC or endogenous glucocorticoids as a result of maternal stress, show increased risk of developing behavioral, endocrine, and metabolic dysregulation. DNA methylation may play a critical role in long-lasting programming of gene regulation underlying these phenotypes. However, peripheral tissues such as blood are often the only accessible source of DNA for epigenetic analyses in humans. Here, we examined the hypothesis that prenatal sGC administration alters DNA methylation signatures in guinea pig offspring hippocampus and whole blood. We compared these signatures across the two tissue types to assess epigenetic biomarkers of common molecular pathways affected by sGC exposure. Guinea pigs were treated with sGC or saline in late gestation. Genome-wide modifications of DNA methylation were analyzed at single nucleotide resolution using reduced representation bisulfite sequencing in juvenile female offspring. Results indicate that there are tissue-specific as well as common methylation signatures of prenatal sGC exposure. Over 90% of the common methylation signatures associated with sGC exposure showed the same directionality of change in methylation. Among differentially methylated genes, 134 were modified in both hippocampus and blood, of which 61 showed methylation changes at identical CpG sites. Gene pathway analyses indicated that prenatal sGC exposure alters the methylation status of gene clusters involved in brain development. These data indicate concordance across tissues of epigenetic programming in response to alterations in glucocorticoid signaling.

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