Detection and analysis of CpG sites with multimodal DNA methylation level distributions and their relationships with SNPs

BackgroundThe epidermal growth factor receptor (EGFR) is a primary target of molecular targeted therapy for lung adenocarcinoma (LUAD). The mechanisms that lead to epigenetic abnormalities of EGFR in LUAD are still unclear. The purpose of our study was to evaluate the abnormal methylation of EGFR CpG sites as potential biomarkers for LUAD.MethodsTo assess the differentially methylation CpG sites of EGFR in LUAD, we used an integrative study of Illumina HumanMethylation450K and RNA-seq data from The Cancer Genome Atlas (TCGA). We evaluated and compared EGFR multiple-omics data to explore the role of CpG sites located in EGFR promoter regions and gene body regions and the association with transcripts, protein expression levels, mutations, and somatic copy number variation. We calculated the correlation coefficients between CpG sites of EGFR and immune infiltration fraction (by MCPcounter and ESTIMATE) and immune-related pathways in LUAD. Finally, we validated the differential methylation of clinically and prognostically relevant CpG sites using quantitative methylation-specific PCR (qMSP).ResultsWe found that the methylation level of many EGFR CpGs in the promoter region was negatively correlated with the transcription level, protein expression, and SCNV, while the methylation at the gene body region was positively correlated with these features. The methylation level of EGFR CpGs in the promoter region was positively correlated with the level of immune infiltration and IFN-γ signature, while the opposite was found for methylation of the gene body region. The qMSP results showed that cg02316066 had a high methylation level, while cg02166842 had a low methylation level in LUAD. There was a high degree of co-methylation between cg02316066 and cg03046247.ConclusionOur data indicate that EGFR is an epigenetic regulator in LUAD acting through DNA methylation. Our research provides a theoretical basis for the further detection of EGFR DNA methylation as a predictive biomarker for LUAD survival and immunotherapy.

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Association between DNA Methylation of the BDNF Promoter Region and Clinical Presentation in Alzheimer's Disease

Dementia and Geriatric Cognitive Disorders Extra ◽

10.1159/000375367 ◽

2015 ◽

Vol 5 (1) ◽

pp. 64-73 ◽

Cited By ~ 41

Author(s):

Tomoyuki Nagata ◽

Nobuyuki Kobayashi ◽

Jumpei Ishii ◽

Shunichiro Shinagawa ◽

Ritsuko Nakayama ◽

...

Keyword(s):

Alzheimer’S Disease ◽

Alzheimer's Disease ◽

Dna Methylation ◽

Methylation Level ◽

Clinical Presentation ◽

Neuropsychological Test ◽

Cpg Sites ◽

The Brain ◽

Normal Controls ◽

Methylation Ratio

Background/Aims: In the present study, we examined whether DNA methylation of the brain-derived neurotrophic factor (BDNF) promoter is associated with the manifestation and clinical presentation of Alzheimer's disease (AD). Methods: Of 20 patients with AD and 20 age-matched normal controls (NCs), the DNA methylation of the BDNF promoter (measured using peripheral blood samples) was completely analyzed in 12 patients with AD and 6 NCs. The resulting methylation levels were compared statistically. Next, we investigated the correlation between the DNA methylation levels and the clinical presentation of AD. Results: The total methylation ratio (in %) of the 20 CpG sites was significantly higher in the AD patients (5.08 ± 5.52%) than in the NCs (2.09 ± 0.81%; p < 0.05). Of the 20 CpG sites, the methylation level at the CpG4 site was significantly higher in the AD subjects than in the NCs (p < 0.05). Moreover, the methylation level was significantly and negatively correlated with some neuropsychological test subscores (registration, recall, and prehension behavior scores; p < 0.05). Conclusion: These results suggest that the DNA methylation of the BDNF promoter may significantly influence the manifestation of AD and might be associated with its neurocognitive presentation.

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DNA methylation patterns of β-globin cluster in β-thalassemia patients

Clinical Epigenetics ◽

10.1186/s13148-020-00987-2 ◽

2020 ◽

Vol 12 (1) ◽

Author(s):

Xiuqin Bao ◽

Yangjin Zuo ◽

Diyu Chen ◽

Cunyou Zhao

Keyword(s):

Dna Methylation ◽

Cord Blood ◽

Methylation Level ◽

Epigenetic Modification ◽

Fetal Hemoglobin ◽

Cpg Sites ◽

Hbf Level ◽

Globin Promoter ◽

Methylation Patterns ◽

Normal Controls

Abstract Background Reactivation of fetal hemoglobin (HbF, α2γ2) holds a therapeutic target for β-thalassemia and sickle cell disease. Although many HbF regulators have been identified, the methylation patterns in β-globin cluster driving the fetal-to-adult hemoglobin switch remains to be determined. Results Here, we evaluated DNA methylation patterns of the β-globin cluster from peripheral bloods of 105 β0/β0 thalassemia patients and 44 normal controls. We also recruited 15 bone marrows and 4 cord blood samples for further evaluation. We identified that the CpG sites in the locus control region (LCR) DNase I hypersensitive site 4 and 3 (HS4-3) regions, and γ- and β-globin promoters displayed hypomethylation in β0/β0-thalassemia patients, especially for the patients with high HbF level, as compared with normal controls. Furthermore, hypomethylations in most of CpG sites of the HS4-3 core regions were also observed in bone marrows (BM) of β0/β0-patients compared with normal controls; and methylation level of γ-globin promoter -50 and + 17 CpG sites showed lower methylation level in patients with high HbF level compared with those with low HbF level and a negative correlation with HbF level among β0-thalassemia patients. Finally, γ-globin promoter + 17 and + 50 CpG sites also displayed significant hypomethylation in cord blood (CB) tissues compared with BM tissues from normal controls. Conclusions Our findings revealed methylation patterns in β-globin cluster associated with β0 thalassemia disease and γ-globin expression, contributed to understand the epigenetic modification in β0 thalassemia patients and provided candidate targets for the therapies of β-hemoglobinopathies.

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Impact of Infrasound on Methylation Status of Genome in Testes of Rats

Journal of Low Frequency Noise Vibration and Active Control ◽

10.1260/026309207781894897 ◽

2007 ◽

Vol 26 (2) ◽

pp. 143-147

Author(s):

Li Rui-Man ◽

Zhuang Zhi-Qiang ◽

Yao Hai-Tao ◽

Pei Zhao-Hui ◽

Chen Jing-Zao

Keyword(s):

Dna Methylation ◽

Methylation Level ◽

Dna Methyltransferase ◽

Methylation Status ◽

Control Groups ◽

Methyltransferase Activity ◽

Dynamic Changes ◽

Time Points ◽

Cpg Sites ◽

Underlying Mechanisms

To probe into the changes induced by infrasonic exposure on the methylation status of the genome in testes and underlying mechanisms, we inspected the percentage of unmethylated CpG sites (an index for DNA methylation level) and DNA methyltransferase activity in testes of SD rats exposed to infrasound of 8Hz at 90dB or 130dB for 1, 7, 14 and 21 days (2h/d). Compared to control groups at the same time points, significantly decreased DNA methylation level and methyltransferase activity were observed in all but experimental groups of ld and 21d at 90dB ( p<0.05). Compared to 90dB groups at the same time points, DNA methylation level and DNA methyltransferase activity at 130dB decreased more significantly ( p<0.05). The patterns of dynamic changes on the percentage of unmethylated CpG sites and methyltransferase activity at 130dB were different from those at 90dB. The results indicated that infrasonic exposure induced epigenetic changes in testes of rats, which depended on parameters of infrasonic exposure.

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Association of Tissue-Specific DNA Methylation Alterations with α-Thalassemia Southeast Asian Deletion

Genetics & Epigenetics ◽

10.1177/1179237x17736107 ◽

2017 ◽

Vol 9 ◽

pp. 1179237X1773610 ◽

Cited By ~ 1

Author(s):

Tanapat Pangeson ◽

Phanchana Sanguansermsri ◽

Torpong Sanguansermsri ◽

Teerapat Seeratanachot ◽

Narutchala Suwanakhon ◽

...

Keyword(s):

Dna Methylation ◽

Methylation Level ◽

Large Deletion ◽

Southeast Asian ◽

Wild Type Allele ◽

Wild Type ◽

Type Allele ◽

Cpg Sites ◽

Potential Marker ◽

Cell Free Fetal Dna

In the wild-type allele, DNA methylation levels of 10 consecutive CpG sites adjacent to the upstream 5′-breakpoint of α-thalassemia Southeast Asian (SEA) deletion are not different between placenta and leukocytes. However, no previous study has reported the map of DNA methylation in the SEA allele. This report aims to show that the SEA mutation is associated with DNA methylation changes, resulting in differential methylation between placenta and leukocytes. Methylation-sensitive high-resolution analysis was used to compare DNA methylation among placenta, leukocytes, and unmethylated control DNA. The result indicates that the DNA methylation between placenta and leukocyte DNA is different and shows that the CpG status of both is not fully unmethylated. Mapping of individual CpG sites was performed by targeted bisulfite sequencing. The DNA methylation level of the 10 consecutive CpG sites was different between placenta and leukocyte DNA. When the 10th CpG of the mutation allele was considered as a hallmark for comparing DNA methylation level, it was totally different from the unmethylated 10th CpG of the wild-type allele. Finally, the distinct DNA methylation patterns between both DNA were extracted. In total, 24 patterns were found in leukocyte samples and 9 patterns were found in placenta samples. This report shows that the large deletion is associated with DNA methylation change. In further studies for clinical application, the distinct DNA methylation pattern might be a potential marker for detecting cell-free fetal DNA.

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Association of Aberrant DNA Methylation Level in the CD4 and JAK-STAT-Pathway-Related Genes with Mastitis Indicator Traits in Chinese Holstein Dairy Cattle

Animals ◽

10.3390/ani12010065 ◽

2021 ◽

Vol 12 (1) ◽

pp. 65

Author(s):

Tahir Usman ◽

Nawab Ali ◽

Yachun Wang ◽

Ying Yu

Keyword(s):

Gene Expression ◽

Dna Methylation ◽

Dairy Cattle ◽

Methylation Level ◽

Promoter Region ◽

Promoter Regions ◽

Cpg Sites ◽

Cell Counts ◽

Aberrant Dna Methylation ◽

Stat Pathway

The present study was designed to evaluate the gene expression and DNA methylation level in the promoter region of the CD4 and the JAK-STAT-pathway-related genes. A total of 24 samples were deployed in the gene expression and 118 samples were used in the DNA methylation study. Student’s t-tests were used to analyze the gene expression and DNA methylation. The evaluation of DNA methylation in promoter regions of JAK2 and STAT5A revealed hypo-methylation levels of CpG sites and higher gene expression in cows diagnosed with mastitis as compared to the healthy control, and vice versa in those with CD4. DNA methylation was negatively correlated with gene expression in JAK2, STAT5A, and CD4 genes. Six, two, and four active transcription factors were identified on the CpG sites in the promoter regions of JAK2, STAT5A, and CD4 genes, respectively. Regarding correlation analysis, the DNA methylation levels of CD4 showed significantly higher positive correlations with somatic cell counts (p < 0.05). Findings of the current study inferred that aberrant DNA methylation in the CpG sites at the 1 kb promoter region in JAK2, STAT5A, and CD4 genes due to mastitis in cows can be used as potential epigenetic markers to estimate bovine mastitis susceptibility in dairy cattle.

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Epigenome-wide analysis of DNA methylation and coronary heart disease: a nested case-control study

eLife ◽

10.7554/elife.68671 ◽

2021 ◽

Vol 10 ◽

Author(s):

Jiahui Si ◽

Songchun Yang ◽

Dianjianyi Sun ◽

Canqing Yu ◽

Yu Guo ◽

...

Keyword(s):

Blood Pressure ◽

Dna Methylation ◽

Methylation Level ◽

Case Control Study ◽

Asian Population ◽

Cpg Sites ◽

Chd Risk ◽

Nested Case Control ◽

Control Study

Background: Identifying environmentally responsive genetic loci where DNA methylation is associated with coronary heart disease (CHD) may reveal novel pathways or therapeutic targets for CHD. We conducted the first prospective epigenome-wide analysis of DNA methylation in relation to incident CHD in the Asian population.Methods: We did a nested case-control study comprising incident CHD cases and 1:1 matched controls who were identified from the 10-year follow-up of the China Kadoorie Biobank. Methylation level of baseline blood leukocyte DNA was measured by Infinium Methylation EPIC BeadChip. We performed the single cytosine-phosphate-guanine (CpG) site association analysis and network approach to identify CHD-associated CpG sites and co-methylation gene module.Results: After quality control, 982 participants (mean age 50.1 years) were retained. Methylation level at 25 CpG sites across the genome was associated with incident CHD (genome-wide false discovery rate [FDR] < 0.05 or module-specific FDR <0.01). One SD increase in methylation level of identified CpGs was associated with differences in CHD risk, ranging from a 47% decrease to a 118% increase. Mediation analyses revealed 28.5% of the excessed CHD risk associated with smoking was mediated by methylation level at the promoter region of ANKS1A gene (P for mediation effect = 0.036). Methylation level at the promoter region of SNX30 was associated with blood pressure and subsequent risk of CHD, with the mediating proportion to be 7.7% (P = 0.003) via systolic blood pressure and 6.4% (P = 0.006) via diastolic blood pressure. Network analysis revealed a co-methylation module associated with CHD.Conclusions: We identified novel blood methylation alterations associated with incident CHD in the Asian population and provided evidence of the possible role of epigenetic regulations in the smoking- and BP-related pathways to CHD risk.Funding: This work was supported by National Natural Science Foundation of China (81390544 and 91846303). The CKB baseline survey and the first re-survey were supported by a grant from the Kadoorie Charitable Foundation in Hong Kong. The long-term follow-up is supported by grants from the UK Wellcome Trust (202922/Z/16/Z, 088158/Z/09/Z, 104085/Z/14/Z), grant (2016YFC0900500, 2016YFC0900501, 2016YFC0900504, 2016YFC1303904) from the National Key and Program of China, and Chinese Ministry of Science and Technology (2011BAI09B01).

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50 DNA METHYLATION PROFILES OF UPSTREAM ELEMENTS OF Oct-3/4 GENE IN IN VITRO FERTILIZATION (IVF) AND SOMATIC CELL NUCLEAR-TRANSFERRED (SCNT) EMBRYOS

Reproduction Fertility and Development ◽

10.1071/rdv19n1ab50 ◽

2007 ◽

Vol 19 (1) ◽

pp. 143

Author(s):

M. Kawasumi ◽

Y. Unno ◽

M. Nishiwaki ◽

K. Matsumoto ◽

M. Anzai ◽

...

Keyword(s):

Dna Methylation ◽

In Vitro Fertilization ◽

Somatic Cell ◽

Methylation Level ◽

Nuclear Transfer ◽

Upstream Region ◽

Sequencing Analysis ◽

Vitro Fertilization ◽

Cpg Sites

Cloning by adult somatic cell nuclear transfer (SCNT) has proven to be successful for the production of clones from many species (Keith 2004 Cytogenet. Genome Res. 105, 285). However, somatic cloning is currently highly inefficient. One of the reasons for this is that SCNT is believed to be associated with epigenetic errors including abnormal DNA methylation of the reconstructed embryo. The Oct-3/4 gene, a member of the POU transcription factor family, is expressed throughout the pre-implantation embryo. Abnormal expression of the Oct-3/4 gene in the nuclear-transferred embryo is either directly or indirectly caused by nuclear transfer and is suggested to be indicative of a general failure to reset the genetic program (Boiani et al. 2002 Genes Dev. 16, 1209). In this study, we investigated the DNA methylation profiles of the Oct-3/4 gene in the genome of SCNT embryos, using bisulfite sequencing analysis. Then, we observed the detailed subcellular localization of Oct-3/4 proteins in SCNT embryos using immunocytochemical (ICC) analysis. Nuclear transfer of cumulus cell nuclei was carried out as previously described (Wakayama et al. 1998 Nature 394, 369). After nuclear transfer, embryos were subsequently cultured in KSOM media to the morula and blastocyst stages. We compared the methylation profiles of 3 transcriptional control elements (distal enhancer, DE; proximal enhancer, PE; and promoter) of the upstream region of the Oct-3/4 gene with the genome of in vitro fertilization (IVF) and SCNT embryos. The methylation rate of CpG sites in the DE and promoter regions of both IVF and SCNT embryos was low at both the morula and the blastocyst stages. What's interesting is that there was a significant difference in the methylation level on CpG sites in the PE element between IVF and SCNT embryos. At the morula stage, the methylation level on CpG sites in the PE element was very low in the IVF embryo and moderately high in the SCNT embryo (0.9&percnt; and 26.3&percnt;). Conversely, at the blastocyst stage, CpG sites in the PE element showed high methylation in the IVF embryo and low methylation in the SCNT embryo (55.2&percnt; and 10.5&percnt;). CpG sites in the PE element were lightly methylated (3.0&percnt;) in the inner cell mass (ICM) of the IVF embryo. This means that the main portion of methylation in the IVF blastocyst embryo occurred at the trophectoderm (TE). On the other hand, in ICM of the SCNT embryo, the methylation level of each embryonic cell was almost the same in the whole blastocyst embryo (9.8&percnt; and 10.5&percnt;). As a result, it is highly possible that the CpG sites in the PE element of ICM were methylated as in the TE. ICC analysis revealed that some SCNT embryos showed aberrant Oct-3/4 expression in the TE. These results indicate that the methylation of CpG sites in the Oct-3/4 PE element may be related to expression of Oct-3/4 in the mouse IVF and SCNT embryos. These differences in methylation level between IVF and SCNT embryos were reflected as abnormal expressions of Oct-3/4 on SCNT embryos. This study was supported by the 21st COE Program of MEST. M.K. is a JSPS Research Fellow and supported by Grant-in Aid for Scientific Research (No. 1751132) of JSPS.

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Genetic analysis of DNA methylation in dyslipidemia: a case-control study

10.21203/rs.3.rs-113737/v1 ◽

2020 ◽

Author(s):

Yi-Tong Ma ◽

Shuai Liu ◽

Yang Li ◽

Xian Wei ◽

Dilare Adi ◽

...

Keyword(s):

Dna Methylation ◽

Methylation Level ◽

Case Control Study ◽

Case Control ◽

Control Group ◽

Cpg Sites ◽

Heart Center ◽

Medical University ◽

The Relationship ◽

Control Study

Abstract In our previous study, we explored the relationship between TBL2 gene DNA methylation and high-low-density lipoproteinemia (Hyper-LDL). However, Hyper-LDL is only one type of dyslipidemia. In order to expand the scope of clinical application, we explored the correlation between DNA methylation of genes related to lipid metabolism and dyslipidemia in this study. This study is a case-control study. A total of 180 samples were included in this study from the Heart Center of the First Affiliated Hospital of Xinjiang Medical University. The BSAS method was used to detect the DNA methylation levels and haplotypes of AMFR, FBXW7, INSIG1, INSIG2, MBTPS1 and GRINA genes. A total of 259 CpG sites and 14 regions were detected. The study found that a total of 24 CpG sites DNA methylation and 20 haplotypes were statistically different. The GRINA gene DNA methylation level in the dyslipidemia group was higher than that in the control group (2.68 vs 2.36, p = 0.04). ttttttttttttcttttttttttt is significant methylation haplotype of GRINA (p=0.017). Through logistics analysis, it is found that GRINA gene DNA methylation is an independent risk factor for dyslipidemia, and the increase of GRINA gene DNA methylation level will increase the prevalence of dyslipidemia.

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Association between the extent of DNA methylation at the CpG sites of HIF3A and parameters of obesity in the general Japanese population

10.1101/2020.11.30.403709 ◽

2020 ◽

Author(s):

Genki Mizuno ◽

Hiroya Yamada ◽

Eiji Munetsuna ◽

Mirai Yamazaki ◽

Yoshitaka Ando ◽

...

Keyword(s):

Body Mass Index ◽

Dna Methylation ◽

Body Mass ◽

Methylation Level ◽

Japanese Population ◽

Public Health Problem ◽

Major Public Health Problem ◽

Transcriptional Responses ◽

General Japanese Population ◽

Cpg Sites

AbstractObesity is a major public health problem worldwide owing to the substantial increase in risk of metabolic diseases. Hypoxia-inducible factors (HIFs) regulate transcriptional responses to hypoxic stress. DNA methylation in the CpG sites of intron 1 of HIF3A is associated with body mass index in the whole blood and adipose tissue. This study investigates the correlation between DNA methylation of HIF3A and parameters of obesity, including thickness of visceral (VAT) and subcutaneous adipose tissues, in the general Japanese population. Participants (220 men and 253 women) who underwent medical examination were enrolled in this cross-sectional study. We used pyrosequencing to quantify DNA methylation (CpG sites of cg16672562, cg22891070, and cg27146050) in HIF3A. DNA methylation of HIF3A was only different in women. Multiple regression analysis showed that DNA methylation level at cg27146050 was associated with thickness of VAT in women. DNA methylation level at cg27146050 also correlated with body mass index and percentage of body fat in women after excluding smokers and non-smokers who quit smoking with the last 5 years. DNA methylation in the CpG site (cg27146050) of HIF3A correlated with parameters of obesity in Japanese women.

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