Heritability and Reversibility of DNA Methylation Induced by in vitro Grafting between Brassica juncea and B. oleracea

DNA methylation mediates organisms’ adaptations to environmental changes in a wide range of species. We investigated if a such a strategy is also adopted by Fusarium graminearum in regulating virulence toward its natural hosts. A virulent strain of this fungus was consecutively sub-cultured for 50 times (once a week) on potato dextrose agar. To assess the effect of subculturing on virulence, wheat seedlings and heads (cv. A416) were inoculated with subcultures (SC) 1, 23, and 50. SC50 was also used to re-infect (three times) wheat heads (SC50×3) to restore virulence. In vitro conidia production, colonies growth and secondary metabolites production were also determined for SC1, SC23, SC50, and SC50×3. Seedling stem base and head assays revealed a virulence decline of all subcultures, whereas virulence was restored in SC50×3. The same trend was observed in conidia production. The DNA isolated from SC50 and SC50×3 was subject to a methylation content-sensitive enzyme and double-digest, restriction-site-associated DNA technique (ddRAD-MCSeEd). DNA methylation analysis indicated 1024 genes, whose methylation levels changed in response to the inoculation on a healthy host after subculturing. Several of these genes are already known to be involved in virulence by functional analysis. These results demonstrate that the physiological shifts following sub-culturing have an impact on genomic DNA methylation levels and suggest that the ddRAD-MCSeEd approach can be an important tool for detecting genes potentially related to fungal virulence.

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Androgenic-Induced Transposable Elements Dependent Sequence Variation in Barley

International Journal of Molecular Sciences ◽

10.3390/ijms22136783 ◽

2021 ◽

Vol 22 (13) ◽

pp. 6783

Author(s):

Renata Orłowska ◽

Katarzyna A. Pachota ◽

Wioletta M. Dynkowska ◽

Agnieszka Niedziela ◽

Piotr T. Bednarek

Keyword(s):

Dna Methylation ◽

Transposable Elements ◽

Dna Sequence ◽

Sequence Variation ◽

Nuclear Dna ◽

Point Mutations ◽

Mobile Elements ◽

Dna Demethylation ◽

Plant Genome

A plant genome usually encompasses different families of transposable elements (TEs) that may constitute up to 85% of nuclear DNA. Under stressful conditions, some of them may activate, leading to sequence variation. In vitro plant regeneration may induce either phenotypic or genetic and epigenetic changes. While DNA methylation alternations might be related, i.e., to the Yang cycle problems, DNA pattern changes, especially DNA demethylation, may activate TEs that could result in point mutations in DNA sequence changes. Thus, TEs have the highest input into sequence variation (SV). A set of barley regenerants were derived via in vitro anther culture. High Performance Liquid Chromatography (RP-HPLC), used to study the global DNA methylation of donor plants and their regenerants, showed that the level of DNA methylation increased in regenerants by 1.45% compared to the donors. The Methyl-Sensitive Transposon Display (MSTD) based on methylation-sensitive Amplified Fragment Length Polymorphism (metAFLP) approach demonstrated that, depending on the selected elements belonging to the TEs family analyzed, varying levels of sequence variation were evaluated. DNA sequence contexts may have a different impact on SV generated by distinct mobile elements belonged to various TE families. Based on the presented study, some of the selected mobile elements contribute differently to TE-related SV. The surrounding context of the TEs DNA sequence is possibly important here, and the study explained some part of SV related to those contexts.

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Cell–cell coupling and DNA methylation abnormal phenotypes in the after-hours mice

Epigenetics & Chromatin ◽

10.1186/s13072-020-00373-5 ◽

2021 ◽

Vol 14 (1) ◽

Author(s):

Federico Tinarelli ◽

Elena Ivanova ◽

Ilaria Colombi ◽

Erica Barini ◽

Edoardo Balzani ◽

...

Keyword(s):

Gene Expression ◽

Dna Methylation ◽

Neuronal Networks ◽

Molecular Mechanisms ◽

Ex Vivo ◽

Neuronal Cultures ◽

Functional Impairments ◽

After Hours ◽

The Impact

Abstract Background DNA methylation has emerged as an important epigenetic regulator of brain processes, including circadian rhythms. However, how DNA methylation intervenes between environmental signals, such as light entrainment, and the transcriptional and translational molecular mechanisms of the cellular clock is currently unknown. Here, we studied the after-hours mice, which have a point mutation in the Fbxl3 gene and a lengthened circadian period. Methods In this study, we used a combination of in vivo, ex vivo and in vitro approaches. We measured retinal responses in Afh animals and we have run reduced representation bisulphite sequencing (RRBS), pyrosequencing and gene expression analysis in a variety of brain tissues ex vivo. In vitro, we used primary neuronal cultures combined to micro electrode array (MEA) technology and gene expression. Results We observed functional impairments in mutant neuronal networks, and a reduction in the retinal responses to light-dependent stimuli. We detected abnormalities in the expression of photoreceptive melanopsin (OPN4). Furthermore, we identified alterations in the DNA methylation pathways throughout the retinohypothalamic tract terminals and links between the transcription factor Rev-Erbα and Fbxl3. Conclusions The results of this study, primarily represent a contribution towards an understanding of electrophysiological and molecular phenotypic responses to external stimuli in the Afh model. Moreover, as DNA methylation has recently emerged as a new regulator of neuronal networks with important consequences for circadian behaviour, we discuss the impact of the Afh mutation on the epigenetic landscape of circadian biology.

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Aberrant DNA methylation of imprinted loci in hepatocellular carcinoma and after in vitro exposure to common risk factors

Clinical Epigenetics ◽

10.1186/s13148-015-0053-9 ◽

2015 ◽

Vol 7 (1) ◽

pp. 15 ◽

Cited By ~ 15

Author(s):

Marie-Pierre Lambert ◽

Pierre-Benoit Ancey ◽

Davide Esposti ◽

Marie-Pierre Cros ◽

Athena Sklias ◽

...

Keyword(s):

Risk Factors ◽

Hepatocellular Carcinoma ◽

Dna Methylation ◽

In Vitro Exposure ◽

Common Risk Factors ◽

Aberrant Dna Methylation

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Epigenetic Modifications During Sex Change Repress Gonadotropin Stimulation of Cyp19a1a in a Teleost Ricefield Eel (Monopterus albus)

Endocrinology ◽

10.1210/en.2012-2220 ◽

2013 ◽

Vol 154 (8) ◽

pp. 2881-2890 ◽

Cited By ~ 54

Author(s):

Yang Zhang ◽

Shen Zhang ◽

Zhixin Liu ◽

Lihong Zhang ◽

Weimin Zhang

Keyword(s):

Dna Methylation ◽

Binding Protein ◽

Sex Change ◽

Gonadal Development ◽

Proximal Region ◽

Histone Deacetylation ◽

Epigenetic Mechanisms ◽

Gonadal Differentiation ◽

Stimulation Of

Abstract In vertebrates, cytochrome P450 aromatase, encoded by cyp19a1, converts androgens to estrogens and plays important roles in gonadal differentiation and development. The present study examines whether epigenetic mechanisms are involved in cyp19a1a expression and subsequent gonadal development in the hermaphroditic ricefield eel. The expression of the ricefield eel cyp19a1a was stimulated by gonadotropin via the cAMP pathway in the ovary but not the ovotestis or testis. The CpG within the cAMP response element (CRE) of the cyp19a1a promoter was hypermethylated in the ovotestis and testis compared with the ovary. The methylation levels of CpG sites around CRE in the distal region (region II) and around steroidogenic factor 1/adrenal 4 binding protein sites and TATA box in the proximal region (region I) were inversely correlated with cyp19a1a expression during the natural sex change from female to male. In vitro DNA methylation decreased the basal and forskolin-induced activities of cyp19a1a promoter. Chromatin immunoprecipitation assays indicated that histone 3 (Lys9) in both regions I and II of the cyp19a1a promoter were deacetylated and trimethylated in the testis, and in contrast to the ovary, phosphorylated CRE-binding protein failed to bind to these regions. Lastly, the DNA methylation inhibitor 5-aza-2′-deoxycytidine reversed the natural sex change of ricefield eels. These results suggested that epigenetic mechanisms involving DNA methylation and histone deacetylation and methylation may abrogate the stimulation of cyp19a1a by gonadotropins in a male-specific fashion. This may be a mechanism widely used to drive natural sex change in teleosts as well as gonadal differentiation in other vertebrates.

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PGC7 suppresses TET3 for protecting DNA methylation

Nucleic Acids Research ◽

10.1093/nar/gkt1261 ◽

2013 ◽

Vol 42 (5) ◽

pp. 2893-2905 ◽

Cited By ~ 39

Author(s):

Chunjing Bian ◽

Xiaochun Yu

Keyword(s):

Dna Methylation ◽

Molecular Mechanism ◽

Biological Process ◽

Cpg Islands ◽

Binding Motifs ◽

Genome Wide Analysis ◽

Dna Motif ◽

Genome Wide

Abstract Ten-eleven translocation (TET) family enzymes convert 5-methylcytosine to 5-hydroxylmethylcytosine. However, the molecular mechanism that regulates this biological process is not clear. Here, we show the evidence that PGC7 (also known as Dppa3 or Stella) interacts with TET2 and TET3 both in vitro and in vivo to suppress the enzymatic activity of TET2 and TET3. Moreover, lacking PGC7 induces the loss of DNA methylation at imprinting loci. Genome-wide analysis of PGC7 reveals a consensus DNA motif that is recognized by PGC7. The CpG islands surrounding the PGC7-binding motifs are hypermethylated. Taken together, our study demonstrates a molecular mechanism by which PGC7 protects DNA methylation from TET family enzyme-dependent oxidation.

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TMOD-02. GENERATION OF A NOVEL MOUSE MODEL FOR BRAIN TUMORS OF THE DNA METHYLATION CLASS “GBM MYCN”

Neuro-Oncology ◽

10.1093/neuonc/noab196.864 ◽

2021 ◽

Vol 23 (Supplement_6) ◽

pp. vi215-vi216

Author(s):

Melanie Schoof ◽

Carolin Göbel ◽

Dörthe Holdhof ◽

Sina Al-Kershi ◽

Ulrich Schüller

Keyword(s):

Dna Methylation ◽

Brain Tumors ◽

Molecular Mechanisms ◽

Treatment Options ◽

Tumor Development ◽

Model Systems ◽

Preclinical Research ◽

Human Gbm

Abstract DNA methylation based classification of brain tumors has revealed a high heterogeneity between tumors and led to the description of multiple distinct subclasses. The increasing subdivision of tumors can help to understand molecular mechanisms of tumor development and to improve therapy if appropriate model systems for preclinical research are available. Multiple recent publications have described a subgroup of pediatric glioblastoma which is clearly separable from other pediatric and adult glioblastoma in its DNA methylation profile (GBM MYCN). Many cases in this group are driven by MYCN amplifications and harbor TP53 mutations. These tumors almost exclusively occur in children and were further described as highly aggressive with a median overall survival of only 14 months. In order to further investigate the biology and treatment options of these tumors, we generated hGFAP-cre::TP53 Fl/Fl ::lsl-MYCN mice. These mice carry a loss of TP53 and show aberrant MYCN expression in neural precursors of the central nervous system. The animals develop large forebrain tumors within the first 80 days of life with 100 % penetrance. These tumors resemble human GBM MYCN tumors histologically and are sensitive to AURKA and ATR inhibitors in vitro. We believe that further characterization of the model and in vivo treatment studies will pave the way to improve treatment of patients with these highly aggressive tumors.

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Abstract 130: Hypercholesterolemia Suppresses Carotid Artery Endothelial NOS3 Expression via Epigenetic Mechanisms

Arteriosclerosis Thrombosis and Vascular Biology ◽

10.1161/atvb.35.suppl_1.130 ◽

2015 ◽

Vol 35 (suppl_1) ◽

Author(s):

Alex Sotolongo ◽

Yi-Zhou Jiang ◽

John Karanian ◽

William Pritchard ◽

Peter Davies

Keyword(s):

Gene Expression ◽

Dna Methylation ◽

Endothelial Cells ◽

Dna Methyltransferase ◽

Control Group ◽

High Cholesterol Diet ◽

Cholesterol Diet ◽

High Cholesterol ◽

High Fat

Objective: One of the first clinically detectable changes in the vasculature during atherogenesis is the accumulation of cholesterol within the vessel wall. Hypercholesterolemia is characterized by dysfunctional endothelial-dependent vessel relaxation and impaired NOS3 function. Since DNA methylation at gene promoter regions strongly suppresses gene expression, we postulated that high-fat/high-cholesterol diet suppresses endothelial NOS3 through promoter DNA methylation. Methods: Domestic male pigs were fed control diet (CD) or isocaloric high fat and high cholesterol diet (HC; 12% fat and 1.5% cholesterol) for 2, 4, 8 or 12 weeks prior to tissue collection. Furthermore, to determine the effects of risk factor withdrawal, an additional group of swine received HC for 12 weeks and then CD for 8 weeks; a control group received HC continuously for 20 weeks. Endothelial cells were harvested from common carotid aorta. In parallel in vitro studies, cultured human aortic endothelial cells (HAEC) were treated with human LDL, GW3956 (LXR agonist) and RG108 (DNA methyltransferase [DNMT] inhibitor). In cells from both sources, DNA methylation at the NOS3 promoter was measured using methylation specific pyro sequencing, and endothelial gene expression was measured using RT PCR. Results: HC diet increased plasma cholesterol level from 75 mg/dl on CD to a plateau of about 540 mg/dl within 2 weeks. Endothelial NOS3 expression was significantly reduced (71±9 % of CD) after 4 weeks of HC, a level sustained at subsequent time points. Withdrawal of HC for 8 weeks did not recover NOS3 expression. After 12-week HC, the NOS3 promoter was hypermethylated. Withdrawal of HC did not reverse NOS3 promoter methylation. In vitro treatment of HAEC with human LDL (200 mg/dl total cholesterol) or GW3956 (5μM) suppressed NOS3 mRNA to 50% and 30% respectively, suggesting that LXR/RXR is involved in suppression of NOS3. Nitric oxide production was consistently suppressed by GW3959. Both could be reversed through inhibition of DNMTs by RG108. Conclusions: DNA methylation and LXR/RXR pathway can mediate the HC-suppression of endothelial NOS3. The study identifies novel pharmaceutical targets in treating endothelial dysfunction. Crosstalk between these pathways is under investigation.

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