Gene Body Regions Research Articles

Integration of transcriptome and DNA methylome analysis reveals the molecular mechanism of taproot yield heterosis in radish (Raphanus sativus L.)

Heterosis, a crucial biological phenomenon, plays a vital role in determining the yield and quality of plants. Radish, an important root vegetable crop, exhibits notable heterosis in terms of root yield and quality. Nevertheless, the specific molecular mechanism behind the formation of heterosis in radish remains unclear. Herein, transcriptome and DNA methylome analyses were performed on F1 hybrids and parental lines. Expression level dominance (ELD) genes and allele-specific expression (ASEG) genes together significantly contribute to heterosis, primarily through energy metabolism and plant hormone signal transduction pathway. Additionally, an increase in the average methylation level in the F1 hybrids was observed compared to the parental lines. Interestingly, a negative correlation was found between the methylation level of differentially expressed genes (DEGs) in gene body regions and their expression levels in NAU-LB and the F1 hybrids. Conversely, a contrasting trend was observed between NAU-YH and the F1 hybrids. Furthermore, when the parental lines and the F1 hybrids were treated with 5-azacytidine, the hybrids were more sensitive to methylation inhibitors than their parents. A significant decrease was observed in root weight and total sugar content in the F1 hybrids compared to the control. Immunolocalization results indicated a significant decrease in the auxin content of the F1 hybrid under 5-azacytidine treatment. Proliferating cell nuclear antigen immunolocalization also revealed significant inhibition of vascular cambium activity in both the hybrids and parental lines. Notably, the expression profiles of a few differentially methylated DEGs including RsSUS1, RsSUC2a, RsIAA7, and RsIAA18, were significantly increased in the root of hybrids compared to their parents, suggesting a potential role for DNA methylation in yield heterosis. Collectively, these findings would provide valuable insight into the molecular mechanism underlying taproot yield heterosis and have the potential to facilitate the genetic improvement of taproot yield and quality in radish breeding programs.

High overall copy number variation burden by genome-wide methylation profiling holds negative prognostic value in surgically treated pancreatic ductal adenocarcinoma

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive type of cancer with an overall 5-year survival of around 10 %. New prognostic tools to stratify patients are needed. Our main aim was to evaluate the prognostic value of overall copy number variation (CNV) burden in surgically treated PDAC. DNA extracted from 108 surgical PDAC specimens was examined to collect data on the genome-wide DNA methylation status of >850,000 CpG sites in promoter, gene body, and enhancer regions (Illumina Infinium Methylation EPIC BeadChip Kit). CNV profiles were obtained and all PDACs were stratified into one of three groups: Low, moderate, or high overall CNV burden. Tumors histologically showing a dominant conventional and/or tubulopapillary pattern in 60 %–100 % and 0–59 % were categorized as Group A and Group B as per Kalimuthu. We also performed targeted next-generation sequencing (NGS) and immunohistochemistry. High overall CNV burden held independent negative prognostic value with poor survival (HR 4.01 (95%CI 1.96–8.19), p = 0.00014) and was more frequent in Group B (p = 0.0003). Most frequent chromosomal arm-level aberrations were gains of 8q (29 %) and 1q (19 %) and losses of 17p (55 %), 18q (43 %), 6q (37 %), 9p (36 %), 6p (26 %), 19p (26 %), and 8p (25 %). Most frequent mutations found were in KRAS (95 %), TP53 (62 %), CDKN2A (24 %), SMAD4 (23 %), ATM (9 %), ARID1A (7 %), RNF43 (7 %), GNAS (6 %), and KDM6A (6 %). Group A PDACs showed more frequently KRAS variants other than Gly12Val and Gly12Asp (p = 0.012). Our data indicate that overall CNV burden using genome-wide methylation profiling may be a useful prognostic tool in surgically treated PDAC. Importantly, our approach, using data from genome-wide methylation profiling for analysis of overall CNV burden, can be performed on formalin-fixed and paraffin embedded PDAC tissues. Future studies should examine the prognostic value of overall CNV burden in unresectable PDAC.

Three-dimensional genome architecture coordinates key regulators of lineage specification in mammary epithelial cells

Although lineage-specific genes have been identified in the mammary gland, little is known about the contribution of the 3D genome organization to gene regulation in the epithelium. Here, we describe the chromatin landscape of the three major epithelial subsets through integration of long- and short-range chromatin interactions, accessibility, histone modifications, and gene expression. While basal genes display exquisite lineage specificity via distal enhancers, luminal-specific genes show widespread promoter priming in basal cells. Cell specificity in luminal progenitors is largely mediated through extensive chromatin interactions with super-enhancers in gene-body regions in addition to interactions with polycomb silencer elements. Moreover, lineage-specific transcription factors appear to be controlled through cell-specific chromatin interactivity. Finally, chromatin accessibility rather than interactivity emerged as a defining feature of the activation of quiescent basal stem cells. This work provides a comprehensive resource for understanding the role of higher-order chromatin interactions in cell-fate specification and differentiation in the adult mouse mammary gland.

Loss of KDM6B epigenetically confers resistance to lipotoxicity in nonalcoholic fatty liver disease-related HCC.

NAFLD caused by abnormalities in hepatic lipid metabolism is associated with an increased risk of developing HCC. The molecular mechanisms underlying the progression of NAFLD-related HCC are not fully understood. We investigated the molecular mechanism and role of KDM6B downregulation in NAFLD-related HCC after the KDM6B gene was identified using microarray analysis as commonly downregulated in mouse NAFLD-related HCC and human nonhepatitis B and nonhepatitis C viral-HCC. The 5-hydroxymethylcytosine levels of KDM6B in HCC cells were determined using glycosylated hydroxymethyl-sensitive PCR. Microarray and chromatin immunoprecipitation analyses using KDM6B-knockout (KO) cells were used to identify KDM6B target genes. Lipotoxicity was assessed using a palmitate-treated cell proliferation assay. Immunohistochemistry was used to evaluate KDM6B expression in human HCC tissues. KDM6B expression levels in HCC cells correlated with the 5-hydroxymethylcytosine levels in the KDM6B gene body region. Gene set enrichment analysis revealed that the lipid metabolism pathway was suppressed in KDM6B-KO cells. KDM6B-KO cells acquired resistance to lipotoxicity (p < 0.01) and downregulated the expression of G0S2, an adipose triglyceride lipase/patatin like phospholipase domain containing 2 (ATGL/PNPLA2) inhibitor, through increased histone H3 lysine-27 trimethylation levels. G0S2 knockdown in KDM6B-expressed HCC cells conferred lipotoxicity resistance, whereas ATGL/PNPLA2 inhibition in the KDM6B-KO cells reduced these effects. Immunohistochemistry revealed that KDM6B expression was decreased in human NAFLD-related HCC tissues (p < 0.001), which was significantly associated with decreased G0S2 expression (p = 0.032). KDM6B-disrupted HCC acquires resistance to lipotoxicity via ATGL/PNPLA2 activation caused by epigenetic downregulation of G0S2 expression. Reduced KDM6B and G0S2 expression levels are common in NAFLD-related HCC. Targeting the KDM6B-G0S2-ATGL/PNPLA2 pathway may be a useful therapeutic strategy for NAFLD-related HCC.

Analysis of Antimalarial Drug Resistance Genes of Plasmodium Falciparum 3d7 to Understand Its Expression: A Bioinformatics Approach

Background Malaria, which poses a threat to half of the world’s population, is one of the most serious infectious diseases. Ethiopia is a nation with a significant malaria burden. In Ethiopia, Plasmodium falciparum accounts for 64% of cases of malaria, with P. vivax causing the remaining instances (34 percent). The disease still claims the lives of countless children globally, mostly in sub-Saharan African nations, and malaria continues to be a significant public health issue in Ethiopia despite various advancements in malaria control measures. The potential for analyzing parasite genetics to support both national and international efforts to eradicate parasites is immense. To analyze regulatory components such as CpG islands, transcription factors (TFs), and their corresponding binding sites (TFBSs) involved in the control of gene expression of Plasmodium falciparum 3D7 isolate drug resistance genes. Results Nine drug resistance-related gene-coding sequences from the NCBI database were examined for this analysis. Only functional genes (protein-coding genes) were taken into consideration. Accordingly, genes affected by Plasmodium falciparum 3D7 drug resistance had 1-6 TSS, and five common candidate motifs (MPfI, MPfII, MPfIII, MPfIV, and MPfV) were found in the promoter prediction by neural network promoter prediction results. According to the study, CpG islands are poorly distributed in both the promoter and gene body regions, which may interfere with the accessibility of the promoter to transcription factors and, ultimately, the production of the genes. Conclusion This in silico analysis of genes encoding Plasmodium falciparum drug resistance-related genes may be useful for enhancing knowledge of the molecular data and supporting the identification of gene regulatory elements in the promoter regions.

A DNA adenine demethylase impairs PRC2-mediated repression of genes marked by a specific chromatin signature

BackgroundThe Fe (II)- and α-ketoglutarate-dependent AlkB family dioxygenases are implicated in nucleotide demethylation. AlkB homolog1 (ALKBH1) is shown to demethylate DNA adenine methylation (6mA) preferentially from single-stranded or unpaired DNA, while its demethylase activity and function in the chromatin context are unclear.ResultsHere, we find that loss-of-function of the rice ALKBH1 gene leads to increased 6mA in the R-loop regions of the genome but has a limited effect on the overall 6mA level. However, in the context of mixed tissues, rather than on individual loci, the ALKBH1 mutation or overexpression mainly affects the expression of genes with a specific combination of chromatin modifications in the body region marked with H3K4me3 and H3K27me3 but depleted of DNA CG methylation. In the similar context of mixed tissues, further analysis reveals that the ALKBH1 protein preferentially binds to genes marked by the chromatin signature and has a function to maintain a high H3K4me3/H3K27me3 ratio by impairing the binding of Polycomb repressive complex 2 (PRC2) to the targets, which is required for both the basal and stress-induced expression of the genes.ConclusionOur findings unravel a function of ALKBH1 to control the balance between the antagonistic histone methylations for gene activity and provide insight into the regulatory mechanism of PRC2-mediated H3K27me3 deposition within the gene body region.

Nucleosome retention by histone chaperones and remodelers occludes pervasive DNA-protein binding.

DNA packaging within chromatin depends on histone chaperones and remodelers that form and position nucleosomes. Cells express multiple such chromatin regulators with overlapping in-vitro activities. Defining specific in-vivo activities requires monitoring histone dynamics during regulator depletion, which has been technically challenging. We have recently generated histone-exchange sensors in Saccharomyces cerevisiae, which we now use to define the contributions of 15 regulators to histone dynamics genome-wide. While replication-independent exchange in unperturbed cells maps to promoters, regulator depletions primarily affected gene bodies. Depletion of Spt6, Spt16 or Chd1 sharply increased nucleosome replacement sequentially at the beginning, middle or end of highly expressed gene bodies. They further triggered re-localization of chaperones to affected gene body regions, which compensated for nucleosome loss during transcription complex passage, but concurred with extensive TF binding in gene bodies. We provide a unified quantitative screen highlighting regulator roles in retaining nucleosome binding during transcription and preserving genomic packaging.

Unraveling Prevalence and Effects of Deleterious Mutations in Maize Elite Lines across Decades of Modern Breeding.

Future breeding is likely to involve the detection and removal of deleterious alleles, which are mutations that negatively affect crop fitness. However, little is known about the prevalence of such mutations and their effects on phenotypic traits in the context of modern crop breeding. To address this, we examined the number and frequency of deleterious mutations in 350 elite maize inbred lines developed over the past few decades in China and the United States. Our findings reveal an accumulation of weakly deleterious mutations and a decrease in strongly deleterious mutations, indicating the dominant effects of genetic drift and purifying selection for the two types of mutations, respectively. We also discovered that slightly deleterious mutations, when at lower frequencies, were more likely to be heterozygous in the developed hybrids. This is consistent with complementation as a potential explanation for heterosis. Subsequently, we found that deleterious mutations accounted for more of the variation in phenotypic traits than nondeleterious mutations with matched minor allele frequencies, especially for traits related to leaf angle and flowering time. Moreover, we detected fewer deleterious mutations in the promoter and gene body regions of differentially expressed genes across breeding eras than in nondifferentially expressed genes. Overall, our results provide a comprehensive assessment of the prevalence and impact of deleterious mutations in modern maize breeding and establish a useful baseline for future maize improvement efforts.

Parallel shift of DNA methylation and gene expression toward the mean in mouse spleen with aging.

Age-associated DNA-methylation drift (AMD) manifests itself in two ways in mammals: global decrease (hypomethylation) and local increase of DNA methylation (hypermethylation). To comprehend the principle behind this bidirectional AMD, we studied methylation states of spatially clustered CpG dinucleotides in mouse splenic DNA using reduced-representation-bisulfite-sequencing (RRBS). The mean methylation levels of whole CpGs declined with age. Promoter-resident CpGs, generally weakly methylated (<5%) in young mice, became hypermethylated in old mice, whereas CpGs in gene-body and intergenic regions, initially moderately (~33%) and extensively (>80%) methylated, respectively, were hypomethylated in the old. Chromosome-wise analysis of methylation revealed that inter-individual heterogeneities increase with age. The density of nearby CpGs was used to classify individual CpGs, which found hypermethylation in CpG-rich regions and hypomethylation in CpG-poor regions. When genomic regions were grouped by methylation level, high-methylation regions tended to become hypomethylated whereas low-methylation regions tended to become hypermethylated, regardless of genomic structure/function. Data analysis revealed that while methylation level and CpG density were interdependent, methylation level was a better predictor of the AMD pattern representing a shift toward the mean. Further analysis of gene-expression data showed a decrease in the expression of highly-expressed genes and an increase in the expression of lowly-expressed genes with age. This shift towards the mean in gene-expression changes was correlated with that of methylation changes, indicating a potential link between the two age-associated changes. Our findings suggest that age-associated hyper- and hypomethylation events are stochastic and attributed to malfunctioning intrinsic mechanisms for methylation maintenance in low- and high-methylation regions, respectively.

SlSAMS1 enhances salt tolerance through regulation DNA methylation of SlGI in tomato

S-adenosylmethionine (SAM), which is synthesized from methionine and ATP catalyzed by S-adenosylmethionine synthetase (SAMS), is an important methyl donor in plants. SAMS and DNA methylation play an important role in the plant response to abiotic stresses. Previous studies have shown that SAMS improves salt tolerance in tomato plants, but it is not clear whether the DNA methylation pathway mediates SAMS-induced salt tolerance. This study confirmed that SlSAMS1-overexpressing plants exhibited improved salt tolerance. Through whole-genome bisulfite sequencing (WGBS) and transcriptome sequencing (RNA-seq) analysis, the study screened the circadian rhythm pathway and identified the gene SlGI in this pathway, which was regulated by SlSAMS1. The gene body region of SlGI, the core gene of the circadian rhythm pathway, was hypermethylated in SlSAMS1-overexpressing plants, and its expression level was significantly increased. Furthermore, the SlGI-overexpressing plants showed higher salt tolerance, less reduction in plant height and fresh weight, lower electrolyte leakage, malondialdehyde and H2O2 content, and higher antioxidant enzyme activity compared to wild type plants. Therefore, SlSAMS1-overexpressing plants regulated significant changes in CHG-type methylation sites of the SlGI gene body and its expression levels, leading to an enhanced salt tolerance of tomato plants.

Epigenomics Analysis of the Suppression Role of SIRT1 via H3K9 Deacetylation in Preadipocyte Differentiation.

Sirtuin 1 (SIRT1) overexpression significantly inhibits lipid deposition during yak intramuscular preadipocyte (YIMA) differentiation; however, the regulatory mechanism remains unknown. We elucidated the role of SIRT1 in YIMA differentiation using lentivirus-mediated downregulation technology and conducted mRNA-seq and ChIP-seq assays using H3K9ac antibodies after SIRT1 overexpression in order to reveal SIRT1 targets during YIMA adipogenesis. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed in order to identify the functional annotation of common genes. In addition, a potential target of SIRT1 was selected to verify its effects on the differentiation and proliferation of YIMAs. SIRT1 interfered with lipid deposition and promoted YIMA differentiation. In total, 143,518 specific peaks were identified after SIRT1 overexpression, where genes associated with downregulation peaks were enriched in transcription, gene expression, lipid-related processes, and classical lipid-related pathways. The H3K9ac signal in the whole genome promoter region (2 kb upstream and downstream of the transcription start site (TSS)) was weakened, and the peaks were distributed across all gene functional regions. Genes that lost signals in their TSS region or gene body region were enriched in both biological processes and pathways associated with lipogenesis. The ChIP-seq results revealed 714 common differential genes in mRNA-seq, which were enriched in "MAPK signaling", "lipid and atherosclerosis", "mTOR signaling", and "FoxO signaling" pathways. A total of 445 genes were downregulated in both their H3K9ac signals and mRNA expression, and one of their most significantly enriched pathways was FoxO signaling. Nine genes (FBP2, FPGT, HSD17B11, KCNJ15, MAP3K20, SLC5A3, TRIM23, ZCCHC10, and ZMYM1) lost the H3K9ac signal in their TSS regions and had low mRNA expression, and three genes (KCNJ15, TGM3, and TRIM54) had low expression but lost their H3K9ac signal in the gene body region. The interference of TRIM23 significantly inhibited fat deposition during preadipocyte differentiation and promoted cell proliferation by increasing S-phase cell numbers. The present study provides new insights into the molecular mechanism of intramuscular fat content deposition and the epigenetic role of SIRT1 in adipocyte differentiation.

Epigenetic Regulation of Subgenomic Gene Expression in Allotetraploid Brassica napus.

The allotetraploid Brasscia napus has now been extensively utilized to reveal the genetic processes involved in hybridization and polyploidization. Here, transcriptome, WGBS, and Chip-Seq sequencing data were obtained to explore the regulatory consequences of DNA methylation and histone modifications on gene expression in B. napus. When compared with diploid parents, the expression levels of 14,266 (about 32%) and 17,054 (about 30%) genes were altered in the An and Cn subgenomes, respectively, and a total of 4982 DEGs were identified in B. napus. Genes with high or no expression in diploid parents often shifted to medium or low expression in B. napus. The number of genes with elevated methylation levels in gene promoters and gene body regions has increased in An and Cn subgenomes. The peak number of H3K4me3 modification increased, while the peak number of H3K27ac and H3K27me3 decreased in An and Cn subgenomes, and more genes that maintained parental histone modifications were identified in Cn subgenome. The differential multiples of DEGs in B. napus were positively correlated with DNA methylation levels in promoters and the gene body, and the differential multiples of these DEGs were also affected by the degree of variation in DNA methylation levels. Further analysis revealed that about 99% of DEGs were of DNA methylation, and about 68% of DEGs were modified by at least two types of DNA methylation and H3K4me3, H3K27ac, and H3K27me3 histone modifications. These results demonstrate that DNA methylation is crucial for gene expression regulation, and different epigenetic modifications have an essential function in regulating the differential expression of genes in B. napus.

The Global Changes of N6-methyldeoxyadenosine in Response to Low Temperature in Arabidopsis thaliana and Rice.

N6-methyldeoxyadenosine (6mA) is a recently discovered DNA modification involved in regulating plant adaptation to abiotic stresses. However, the mechanisms and changes of 6mA under cold stress in plants are not yet fully understood. Here, we conducted a genome-wide analysis of 6mA and observed that 6mA peaks were predominantly present within the gene body regions under both normal and cold conditions. In addition, the global level of 6mA increased both in Arabidopsis and rice after the cold treatment. The genes that exhibited an up-methylation showed enrichment in various biological processes, whereas there was no significant enrichment observed among the down-methylated genes. The association analysis revealed a positive correlation between the 6mA level and the gene expression level. Joint analysis of the 6mA methylome and transcriptome of Arabidopsis and rice unraveled that fluctuations in 6mA levels caused by cold exposure were not correlated to changes in transcript levels. Furthermore, we discovered that orthologous genes modified by 6mA showed high expression levels; however, only a minor amount of differentially 6mA-methylated orthologous genes were shared between Arabidopsis and rice under low-temperature conditions. In conclusion, our study provides information on the role of 6mA in response to cold stress and reveals its potential for regulating the expression of stress-related genes.

METB-10. EXPLAINABLE ARTIFICIAL INTELLIGENCE REVEALS DNA METHYLATION PATTERNS UNDERLYING BRAIN TUMOR CLASSIFICATION

Abstract Over 100 different molecular classes of brain tumors are recognized across histopathological grades and age groups. Their precise diagnosis is crucial for prognostication and appropriate treatment decisions, particularly in pediatric patients. Recently, a random forest-based classifier trained on genome-wide DNA methylation profiles has been developed to guide the accurate classification of these tumors. Over recent years, this classifier has been applied to tens-of-thousands of patient tumors all over the world. Clinical and basic neuro-oncology research would benefit greatly from a detailed understanding of the underlying artificial intelligence decision process. However, why the algorithm arrives at a certain prediction remains unclear. Here, we develop an interpretable framework to explain the model’s decisions and to uncover DNA methylation patterns distinguishing different tumor classes. Using this approach, we demonstrate both global and local differences across epigenetic landscapes. For instance, the model frequently uses DNA methylation probes in CpG islands to distinguish IDH mutant gliomas from other tumor classes. Tumor classes associated with global hypomethylation are distinguished by probes located in broad heterochromatic domains. At the level of individual genes, we find hundreds of promoter and gene body regions specifically methylated or unmethylated in specific classes. In general, we detect a high degree of redundant information in the dataset, with multiple genes across the genome distinguishing individual tumor classes, and many CpG probes representing each gene. These findings explain the technical robustness of the array-based classifier, and provide a starting point for the design of more compact point of care assays. The results of our approach are provided to the research community via an interactive web interface. We hope that this resource will help to build up trust in the machine learning-based classifier in clinical settings, foster biomarker discovery, and enable cancer epigenome research in the context of the central nervous system tumors.

The Methylation Analysis of the Glucose-Dependent Insulinotropic Polypeptide Receptor (GIPR) Locus in GH-Secreting Pituitary Adenomas.

The glucose-dependent insulinotropic polypeptide receptor (GIPR) is aberrantly expressed in about one-third of GH-secreting pituitary adenomas (GH-PAs) and has been associated with a paradoxical increase of GH after a glucose load. The reason for such an overexpression has not yet been clarified. In this work, we aimed to evaluate whether locus-specific changes in DNA methylation patterns could contribute to this phenomenon. By cloning bisulfite-sequencing PCR, we compared the methylation pattern of the GIPR locus in GIPR-positive (GIPR+) and GIPR-negative (GIPR-) GH-PAs. Then, to assess the correlation between Gipr expression and locus methylation, we induced global DNA methylation changes by treating the lactosomatotroph GH3 cells with 5-aza-2'-deoxycytidine. Differences in methylation levels were observed between GIPR+ and GIPR- GH-PAs, both within the promoter (31.9% vs. 68.2%, p < 0.05) and at two gene body regions (GB_1 20.7% vs. 9.1%; GB_2 51.2% vs. 65.8%, p < 0.05). GH3 cells treated with 5-aza-2'-deoxycytidine showed a ~75% reduction in Gipr steady-state level, possibly associated with the observed decrease in CpGs methylation. These results indicate that epigenetic regulation affects GIPR expression in GH-PAs, even though this possibly represents only a part of a much more complex regulatory mechanism.

Impacts of DNA methylases and demethylases on the methylation and expression of Arabidopsis ethylene signal pathway genes.

Arabidopsis ethylene (ET) signal pathway plays important roles in various aspects. Cytosine DNA methylation is significant in controlling gene expression in plants. Here, we analyzed the bisulfite sequencing and mRNA sequencing data from Arabidopsis (de)methylase mutants met1, cmt3, drm1/2, ddm1, ros1-4, and rdd to investigate how DNA (de)methylases influence the DNA methylation and expression of Arabidopsis ET pathway genes. At least 32 genes are found to involved in Arabidopsis ET pathway by text mining. Among them, 14 genes are unmethylated or methylated with very low levels. ACS6 and ACS9 are conspicuously methylated within their upstream regions. The other 16 genes are predominantly methylated at the CG sites within gene body regions in wild-type plants, and mutation of MET1 resulted in almost entire elimination of the CG methylations. In addition, CG methylations within some genes are jointly maintained by MET1 and other (de)methylases. Analyses of mRNA-seq data indicated that some ET pathway genes were differentially expressed between wild-type and diverse mutants. PDF1.2, the marker gene of ET signal pathway, was found being regulated indirectly by the methylases. Eighty-two transposable elements (TEs) were identified to be associated to 15 ET pathway genes. ACS11 is found located in a heterochromatin region that contains 57 TEs, indicating its specific expression and regulation. Together, our results suggest that DNA (de)methylases are implicated in the regulation of CG methylation within gene body regions and transcriptional activity of some ET pathway genes and that maintenance of normal CG methylation is essential for ET pathway in Arabidopsis.

Epigenetic modifier alpha-ketoglutarate modulates aberrant gene body methylation and hydroxymethylation marks in diabetic heart

BackgroundDiabetic cardiomyopathy (DCM) is a leading cause of death in diabetic patients. Hyperglycemic myocardial microenvironment significantly alters chromatin architecture and the transcriptome, resulting in aberrant activation of signaling pathways in a diabetic heart. Epigenetic marks play vital roles in transcriptional reprogramming during the development of DCM. The current study is aimed to profile genome-wide DNA (hydroxy)methylation patterns in the hearts of control and streptozotocin (STZ)-induced diabetic rats and decipher the effect of modulation of DNA methylation by alpha-ketoglutarate (AKG), a TET enzyme cofactor, on the progression of DCM.MethodsDiabetes was induced in male adult Wistar rats with an intraperitoneal injection of STZ. Diabetic and vehicle control animals were randomly divided into groups with/without AKG treatment. Cardiac function was monitored by performing cardiac catheterization. Global methylation (5mC) and hydroxymethylation (5hmC) patterns were mapped in the Left ventricular tissue of control and diabetic rats with the help of an enrichment-based (h)MEDIP-sequencing technique by using antibodies specific for 5mC and 5hmC. Sequencing data were validated by performing (h)MEDIP-qPCR analysis at the gene-specific level, and gene expression was analyzed by qPCR. The mRNA and protein expression of enzymes involved in the DNA methylation and demethylation cycle were analyzed by qPCR and western blotting. Global 5mC and 5hmC levels were also assessed in high glucose-treated DNMT3B knockdown H9c2 cells.ResultsWe found the increased expression of DNMT3B, MBD2, and MeCP2 with a concomitant accumulation of 5mC and 5hmC, specifically in gene body regions of diabetic rat hearts compared to the control. Calcium signaling was the most significantly affected pathway by cytosine modifications in the diabetic heart. Additionally, hypermethylated gene body regions were associated with Rap1, apelin, and phosphatidyl inositol signaling, while metabolic pathways were most affected by hyperhydroxymethylation. AKG supplementation in diabetic rats reversed aberrant methylation patterns and restored cardiac function. Hyperglycemia also increased 5mC and 5hmC levels in H9c2 cells, which was normalized by DNMT3B knockdown or AKG supplementation.ConclusionThis study demonstrates that reverting hyperglycemic damage to cardiac tissue might be possible by erasing adverse epigenetic signatures by supplementing epigenetic modulators such as AKG along with an existing antidiabetic treatment regimen.

Abstract NG06: Investigating biological mediators of social and environmental risk factors on cancer health disparities

Abstract NG06: Investigating biological mediators of social and environmental risk factors on cancer health disparities

Integrated analysis of genome-wide gene expression and DNA methylation profiles reveals candidate genes in ovary endometriosis.

The incidence of endometriosis (EMs), a common disease in gynecology, has increased over the years. Women suffer from the symptoms caused by EMs, such as chronic pelvic pain, dysmenorrhea, and infertility. However, the etiology and pathophysiology of EMs remain unclear. This study aimed to identify candidate genes of endometriosis through integrated analysis of genome-wide gene expression and DNA methylation profiles. Eutopic and ectopic endometrial tissues were collected from patients who were diagnosed as ovarian EMs. Genome-wide methylation profiling identified 17551 differentially methylated loci, with 9777 hypermethylated and 7774 hypomethylated loci. Differentially methylated loci were mainly concentrated in the gene body and intergenic regions. Genome-wide gene expression profiling identified 1837 differentially expressed genes (DEGs), with 1079 genes upregulated and 758 downregulated in ectopic groups. Integrated analysis revealed that DNA methylation was negatively correlated to gene expression in most genomic regions, such as exon, 3'UTR, 5'UTR, and promoter. We also identified promoter-related (53 downregulated and 113 upregulated) and enhancer-related DMGs (212 downregulated and 232 upregulated), which were significantly correlated to the gene expression. Further validation of the top-ranked genes belonging to differentially methylated genes (DMGs) and DEGs revealed that TMEM184A, GREM2, SFN, KIR3DX1, HPGD, ESR1, BST2, PIK3CG and RNASE1 were significant candidate genes in ovarian endometriosis. Our study revealed the significance of DNA methylation in the gene expression in ovary endometriosis, which provides new insights and a molecular foundation for understanding the underlying mechanisms of endometriosis.

The pattern of alternative splicing and DNA methylation alteration and their interaction in linseed (Linum usitatissimum L.) response to repeated drought stresses

BackgroundDrought stress has significantly hampered agricultural productivity worldwide and can also result in modifications to DNA methylation levels. However, the dynamics of DNA methylation and its association with the changes in gene transcription and alternative splicing (AS) under drought stress are unknown in linseed, which is frequently cultivated in arid and semiarid regions.ResultsWe analysed AS events and DNA methylation patterns in drought-tolerant (Z141) and drought-sensitive (NY-17) linseed under drought stress (DS) and repeated drought stress (RD) treatments. We found that the number of intron-retention (IR) and alternative 3’ splice site (Alt3’SS) events were significantly higher in Z141 and NY-17 under drought stress. We found that the linseed response to the DS treatment was mainly regulated by transcription, while the response to the RD treatment was coregulated by transcription and AS. Whole genome-wide DNA methylation analysis revealed that drought stress caused an increase in the overall methylation level of linseed. Although we did not observe any correlation between differentially methylated genes (DMGs) and differentially spliced genes (DSGs) in this study, we found that the DSGs whose gene body region was hypermethylated in Z141 and hypomethylated in NY-17 were enriched in abiotic stress response Gene Ontology (GO) terms. This finding implies that gene body methylation plays an important role in AS regulation in some specific genes.ConclusionOur study is the first comprehensive genome-wide analysis of the relationship between linseed methylation changes and AS under drought and repeated drought stress. Our study revealed different interaction patterns between differentially expressed genes (DEGs) and DSGs under DS and RD treatments and differences between methylation and AS regulation in drought-tolerant and drought-sensitive linseed varieties. The findings will probably be of interest in the future. Our results provide interesting insights into the association between gene expression, AS, and DNA methylation in linseed under drought stress. Differences in these associations may account for the differences in linseed drought tolerance.