CHG Contexts Research Articles

Comprehensive analysis of epigenetic modifications in alfalfa under cadmium stress

Epigenetics plays an important role in plant growth and development and in environmental adaptation. Alfalfa, an important forage crop, is rich in nutrients. However, little is known about the molecular regulatory mechanisms underlying the response of alfalfa to cadmium (Cd) stress. Here, we performed DNA methylation (5mC), RNA methylation (m6A) and transcriptomic sequencing analyses of alfalfa roots under Cd stress. Whole-genome methylation sequencing and transcriptomic sequencing revealed that Cd stress reduced DNA methylation levels. Moreover, a reduced 5mC methylation level was associated with decreased expression of several DNA methyltransferase genes. Compared with those under normal (CK) conditions, the m6A modification levels under Cd stress were greater and were positively correlated with gene expression in alfalfa roots. We also found a negative correlation between the 5mC level and the m6A level, especially in CG and CHG contexts. In yeast, the overexpression of MsNARMP5 (natural resistance-associated macrophage protein) and MsPCR2 (plant cadmium resistance 2), which are modified by 5mC or m6A, significantly increased Cd stress tolerance. These results provide candidate genes for future studies on the mechanism of Cd stress tolerance in alfalfa roots and valuable information for studying heavy metal stress in alfalfa breeding.

DNA Methylation Participates in Drought Stress Memory and Response to Drought in Medicago ruthenica.

Background: Drought is currently a global environmental problem, which inhibits plant growth and development and seriously restricts crop yields. Many plants exposed to drought stress can generate stress memory, which provides some advantages for resisting recurrent drought. DNA methylation is a mechanism involved in stress memory formation, and many plants can alter methylation levels to form stress memories; however, it remains unclear whether Medicago ruthenica exhibits drought stress memory, as the epigenetic molecular mechanisms underlying this process have not been described in this species. Methods: We conducted methylome and transcriptome sequencing to identify gene methylation and expression changes in plants with a history of two drought stress exposures. Results: Methylation analysis showed that drought stress resulted in an approximately 4.41% decrease in M. ruthenica genome methylation levels. The highest methylation levels were in CG dinucleotide contexts, followed by CHG contexts, with CHH contexts having the lowest levels. Analysis of associations between methylation and transcript levels showed that most DNA methylation was negatively correlated with gene expression except methylation within CHH motifs in gene promoter regions. Genes were divided into four categories according to the relationship between methylation and gene expression; the up-regulation of hypo-methylated gene expression accounted for the vast majority (692 genes) and included genes encoding factors key for abscisic acid (ABA) and proline synthesis. The hypo-methylation of the promoter and body regions of these two gene groups induced increased gene transcription levels. Conclusions: In conclusion, DNA methylation may contribute to drought stress memory formation and maintenance in M. ruthenica by increasing the transcription levels of genes key for ABA and proline biosynthesis.

Endogenous viral elements are targeted by RNA silencing pathways in banana.

Endogenous banana streak virus (eBSV) integrants derived from three distinct species, present in Musa balbisiana (B) but not Musa acuminata (A) banana genomes are able to reconstitute functional episomal viruses causing banana streak disease in interspecific triploid AAB banana hybrids but not in the diploid (BB) parent line, which harbours identical eBSV loci. Here, we investigated the regulation of these eBSV. In-depth characterization of siRNAs, transcripts and methylation derived from eBSV using Illumina and bisulfite sequencing were carried out on eBSV-free Musa acuminata AAA plants and BB or AAB banana plants with eBSV. eBSV loci produce low-abundance transcripts covering most of the viral sequence and generate predominantly 24-nt siRNAs. siRNA accumulation is restricted to duplicated and inverted viral sequences present in eBSV. Both siRNA-accumulating and nonaccumulating sequences of eBSV in BB plants are heavily methylated in all three CG, CHG and CHH contexts. Our data suggest that eBSVs are controlled at the epigenetic level in BB diploids. This regulation not only prevents their awakening and systemic infection of the plant but is also probably involved in the inherent resistance of the BB plants to mealybug-transmitted viral infection. These findings are thus of relevance to other plant resources hosting integrated viruses.

Exploring changes in social spider DNA methylation profiles in all cytosine contexts following infection

Living at high density and with low genetic diversity are factors that should both increase the susceptibility of organisms to disease. Therefore, group living organisms, especially those that are inbred, should be especially vulnerable to infection and therefore have particular strategies to cope with infection. Phenotypic plasticity, underpinned by epigenetic changes, could allow group living organisms to rapidly respond to infection challenges. To explore the potential role of epigenetic modifications in the immune response to a group-living species with low genetic diversity, we compared the genome-wide DNA methylation profiles of five colonies of social spiders (Stegodyphus dumicola) in their natural habitat in Namibia at the point just before they succumbed to infection to a point at least six months previously where they were presumably healthier. We found increases in genome- and chromosome-wide methylation levels in the CpG, CHG, and CHH contexts, although the genome-wide changes were not clearly different from zero. These changes were most prominent in the CHG context, especially at a narrow region of chromosome 13, hinting at an as-of-yet unsuspected role of this DNA methylation context in phenotypic plasticity. However, there were few clear patterns of differential methylation at the base level, and genes with a known immune function in spiders had mean methylation changes close to zero. Our results suggest that DNA methylation may change with infection at large genomic scales, but that this type of epigenetic change is not necessarily integral to the immune response of social spiders.

Difference in DNA methylation pattern and expression of active ingredients between garden ginseng and ginseng under forest

Panax ginseng is widely used for its medicinal value. With the decline in wild ginseng resources, garden ginseng (GG) and ginseng under forest (FG) have become the main sources. Their different growth conditions and ages may account for the differences in epigenetics. The genomic DNA of Panax ginseng was digested using the methylation-insensitive enzyme MseⅠ combined with the methylation-sensitive enzymes AciⅠ, PstⅠ, and EcoT22Ⅰ. Then, a sequencing library of CG, CHG, and CHH methylation contexts was constructed and sequenced through amplicon sequencing. Differences in DNA methylation were observed using the methylation context-sensitive enzyme ddRAD (MCSeEd), revealing eight differentially methylated regions (DMRs). The bisulfite sequencing technology revealed that the methylation level of GG was higher than that of FG. The Plant ISOform Sequencing Database was used to predict the regulatory function of CpG island–coding region so as to estimate the relationship between methylation and effective components. The quantitative real-time polymerase chain reaction (qPCR) showed that the expression of functional genes in FG was higher than that in GG. These studies showed that the increase in GG methylation level was associated with the decrease in gene expression, providing a new perspective on epigenetic differences and gene expression in ginseng.

Heat stress induced cytosine methylation in the coding region of Rubisco activase (Rca) reveals its genotype-specific expression in contrasting wheat genotypes

Rising environmental temperature has become a major concern in global wheat production since it has critical effects on plant growth,development and yield. Elevated temperatures have become. Further understanding of the processes behind the development of heattolerance is necessary for significant agricultural crops. Cytosine methylation in DNA plays an essential function in epigenetic regulation of gene expression at various developmental stages and environmental stress in plants. In this study, we report the comparative analysis of cytosine methylation in the coding region of the Rubisco activase (Rca) gene in heat tolerant (RAJ3765) and heat susceptible (HUW510)genotypes of wheat in four growth stages under control and heat-stressed conditions in CG, CHG and CGG contexts. We found that the overall 5-mC increased due to heat stress in HUW510 during tillering (25%), boot stage (25%), heading (48%) and anthesis (50%) as compared to RAJ3765 during tillering (21%), boot stage (4%), heading (4%) and anthesis (11%). Additionally, gene expression profiling by using qPCR at the different plant growth stages also showed the decline in the gene expression in the leaf samples in both the genotypes due to heat stress, with a minimum in the susceptible genotype at the anthesis stage. The study will increase the knowledge on the molecular regulation of photosynthetic pathways by cytosine methylation which would assist in selection and manipulation of heat tolerance in wheat.

Sulfonamide-induced DNA hypomethylation disturbed sugar metabolism in rice (Oryza sativa L.)

DNA methylation is well-accepted as a bridge to unravel the complex interplay between genome and environmental exposures, and its alteration regulated the cellular metabolic responses towards pollutants. However, the mechanism underlying site-specific aberrant DNA methylation and metabolic disorders under pollutant stresses remained elusive. Herein, the multilevel omics interferences of sulfonamides (i.e., sulfadiazine and sulfamerazine), a group of antibiotics pervasive in farmland soils, towards rice in 14 days of 1 mg/L hydroponic exposure were systematically evaluated. Metabolome and transcriptome analyses showed that 57.1–71.4 % of mono- and disaccharides were accumulated, and the differentially expressed genes were involved in the promotion of sugar hydrolysis, as well as the detoxification of sulfonamides. Most differentially methylated regions (DMRs) were hypomethylated ones (accounting for 87–95 %), and 92 % of which were located in the CHH context (H = A, C, or T base). KEGG enrichment analysis revealed that CHH-DMRs in the promoter regions were enriched in sugar metabolism. To reveal the significant hypomethylation of CHH, multi-spectroscopic and thermodynamic approaches, combined with molecular simulation were conducted to investigate the molecular interaction between sulfonamides and DNA in different sequence contexts, and the result demonstrated that sulfonamides would insert into the minor grooves of DNA, and exhibited a stronger affinity with the CHH contexts of DNA compared to CG or CHG contexts. Computational modeling of DNA 3D structures further confirmed that the binding led to a pitch increase of 0.1 Å and a 3.8° decrease in the twist angle of DNA in the CHH context. This specific interaction and the downregulation of methyltransferase CMT2 (log2FC = –4.04) inhibited the DNA methylation. These results indicated that DNA methylation-based assessment was useful for metabolic toxicity prediction and health risk assessment.

Increased DNA methylation contributes to the early ripening of pear fruits during domestication and improvement

BackgroundDNA methylation is an essential epigenetic modification. However, its contribution to trait changes and diversity in the domestication of perennial fruit trees remains unknown.ResultsHere, we investigate the variation in DNA methylation during pear domestication and improvement using whole-genome bisulfite sequencing in 41 pear accessions. Contrary to the significant decrease during rice domestication, we detect a global increase in DNA methylation during pear domestication and improvement. We find this specific increase in pear is significantly correlated with the downregulation of Demeter-like1 (DML1, encoding DNA demethylase) due to human selection. We identify a total of 5591 differentially methylated regions (DMRs). Methylation in the CG and CHG contexts undergoes co-evolution during pear domestication and improvement. DMRs have higher genetic diversity than selection sweep regions, especially in the introns. Approximately 97% of DMRs are not associated with any SNPs, and these DMRs are associated with starch and sucrose metabolism and phenylpropanoid biosynthesis. We also perform correlation analysis between DNA methylation and gene expression. We find genes close to the hypermethylated DMRs that are significantly associated with fruit ripening. We further verify the function of a hyper-DMR-associated gene, CAMTA2, and demonstrate that overexpression of CAMTA2 in tomato and pear callus inhibits fruit ripening.ConclusionsOur study describes a specific pattern of DNA methylation in the domestication and improvement of a perennial pear tree and suggests that increased DNA methylation plays an essential role in the early ripening of pear fruits.

Genomic and Epigenomic Changes in the Progeny of Cold-Stressed Arabidopsis thaliana Plants.

Plants are continuously exposed to various environmental stresses. Because they can not escape stress, they have to develop mechanisms of remembering stress exposures somatically and passing it to the progeny. We studied the Arabidopsis thaliana ecotype Columbia plants exposed to cold stress for 25 continuous generations. Our study revealed that multigenerational exposure to cold stress resulted in the changes in the genome and epigenome (DNA methylation) across generations. Main changes in the progeny were due to the high frequency of genetic mutations rather than epigenetic changes; the difference was primarily in single nucleotide substitutions and deletions. The progeny of cold-stressed plants exhibited the higher rate of missense non-synonymous mutations as compared to the progeny of control plants. At the same time, epigenetic changes were more common in the CHG (C = cytosine, H = cytosine, adenine or thymine, G = guanine) and CHH contexts and favored hypomethylation. There was an increase in the frequency of C to T (thymine) transitions at the CHH positions in the progeny of cold stressed plants; because this type of mutations is often due to the deamination of the methylated cytosines, it can be hypothesized that environment-induced changes in methylation contribute to mutagenesis and may be to microevolution processes and that RNA-dependent DNA methylation plays a crucial role. Our work supports the existence of heritable stress response in plants and demonstrates that genetic changes prevail.

Hypermethylation in the promoter regions of flavonoid pathway genes is associated with skin color fading during 'Daihong' apple fruit development.

Apple fruit skin color fading is not well understood although the molecular mechanism of skin color formation is well known. The red-fleshed apple cultivar 'Daihong' (DH) exhibited fading skin color during fruit development despite having a heterozygous R6 allele but lacking Red-TE for red fruit skin. In this study, transcriptomic analysis revealed the expression level of MdMYB10 increased with fruit development whereas reduced expression levels of MdMYBPA1, MdCHS, MdANS, MdUFGT, MdLAR, and MdANR were observed, consistent with decreased levels of chalcone, anthocyanin, catechin, epicatechin, and procyanidin B2. Whole-genome bisulfite sequencing (WGBS) indicated a global gain in cytosine methylation levels and increased methylation in 5' and 3' flanking regions of genes and transposable elements (TEs), and in TE bodies in all CG, CHG and CHH contexts, especially the mCHH context, during fruit development. The increased DNA methylation was attributed to reduced expression levels of DNA demethylase genes, including MdDME1, MdROS1, and MdROS2. Association analysis revealed a significant negative correlation between promoter methylation levels of MdCHS, MdCHI, MdMYBPA1, and their respective transcript levels, as well as a negative correlation between promoter methylation levels of MdCHS, MdCHI, MdANR, and MdFLS, and the content of chalcones, naringenin-7-glucoside, epicatechin, and quercetin. Treatment with the DNA demethylation agent 5-aza-2'-deoxycytidine verified the negative correlation between DNA methylation and gene expression within the flavonoid pathway. These findings suggest that hypermethylation in promoter regions of genes of the flavonoid biosynthesis pathway is associated with the reduction of gene expression and flavonoid content, and fruit skin color fading during DH apple development.

Differentially methylated genes involved in reproduction and ploidy levels in recent diploidized and tetraploidized Eragrostis curvula genotypes

Epigenetics studies changes in gene activity without changes in the DNA sequence. Methylation is an epigenetic mechanism important in many pathways, such as biotic and abiotic stresses, cell division, and reproduction. Eragrostis curvula is a grass species reproducing by apomixis, a clonal reproduction by seeds. This work employed the MCSeEd technique to identify deferentially methylated positions, regions, and genes in the CG, CHG, and CHH contexts in E. curvula genotypes with similar genomic backgrounds but with different reproductive modes and ploidy levels. In this way, we focused the analysis on the cvs. Tanganyika INTA (4x, apomictic), Victoria (2x, sexual), and Bahiense (4x, apomictic). Victoria was obtained from the diploidization of Tanganyika INTA, while Bahiense was produced from the tetraploidization of Victoria. This study showed that polyploid/apomictic genotypes had more differentially methylated positions and regions than the diploid sexual ones. Interestingly, it was possible to observe fewer differentially methylated positions and regions in CG than in the other contexts, meaning CG methylation is conserved across the genotypes regardless of the ploidy level and reproductive mode. In the comparisons between sexual and apomictic genotypes, we identified differentially methylated genes involved in the reproductive pathways, specifically in meiosis, cell division, and fertilization. Another interesting observation was that several differentially methylated genes between the diploid and the original tetraploid genotype recovered their methylation status after tetraploidization, suggesting that methylation is an important mechanism involved in reproduction and ploidy changes.

DNA methylome analysis provides insights into gene regulatory mechanism for better performance of rice under fluctuating environmental conditions: epigenomics of adaptive plasticity.

Roots play an important role in adaptive plasticity of rice under dry/direct-sown conditions. However, hypomethylation of genes in leaves (resulting in up-regulated expression) complements the adaptive plasticity of Nagina-22 under DSR conditions. Rice is generally cultivated by transplanting which requires plenty of water for irrigation. Such a practice makes rice cultivation a challenging task under global climate change and reducing water availability. However, dry-seeded/direct-sown rice (DSR) has emerged as a resource-saving alternative to transplanted rice (TPR). Though some of the well-adapted local cultivars are used for DSR, only limited success has been achieved in developing DSR varieties mainly because of a limited knowledge of adaptability of rice under fluctuating environmental conditions. Based on better morpho-physiological and agronomic performance of Nagina-22 (N-22) under DSR conditions, N-22 and IR-64 were grown by transplanting and direct-sowing and used for whole genome methylome analysis to unravel the epigenetic basis of adaptive plasticity of rice. Comparative methylome and transcriptome analyses indicated a large number (4078) of genes regulated through DNA methylation/demethylation in N-22 under DSR conditions. Gene × environment interactions play important roles in adaptive plasticity of rice under direct-sown conditions. While genes for pectinesterase, LRK10, C2H2 zinc-finger protein, splicing factor, transposable elements, and some of the unannotated proteins were hypermethylated, the genes for regulation of transcription, protein phosphorylation, etc. were hypomethylated in CG context in the root of N-22, which played important roles in providing adaptive plasticity to N-22 under DSR conditions. Hypomethylation leading to up-regulation of gene expression in the leaf complements the adaptive plasticity of N-22 under DSR conditions. Moreover, differential post-translational modification of proteins and chromatin assembly/disassembly through DNA methylation in CHG context modulate adaptive plasticity of N-22. These findings would help developing DSR cultivars for increased water-productivity and ecological efficiency.

Integrative transcriptome and whole-genome bisulfite sequencing analyses of a temperature-sensitive albino tea plant cultivar.

Green tea made from albino buds and leaves has a strong umami taste and aroma. The cultivar 'Zhonghuang 2' (ZH2, Camellia sinensis) is a natural mutant with young shoots that are yellow in spring and green or yellow-green in summer. However, the mechanism of leaf color change remains unclear. Here, we found that young shoots of ZH2 were yellow at low temperature (LT) and green at high temperature (HT), indicating that ZH2 is a temperature-sensitive cultivar. Transmission electron microscopy analysis showed that the grana in the chloroplasts of young shoots grown at LT were poorly stacked, which caused a lack of photoreactions and chlorophyll. RNA-seq results showed 1279 genes differentially expressed in the young shoots grown at LT compared with those at HT, including genes related to cytochrome synthesis, chloroplast development, photosynthesis, and DNA methylation. A whole-genome bisulfite sequencing assay revealed that the dynamics of DNA methylation levels in the CG, CHG, and CHH contexts decreased under LT, and the change was most obvious in the CHH context. Furthermore, 72 genes showed significant changes in both expression and DNA methylation levels, and most of them were related to cytochrome synthesis, chloroplast development, photosynthesis, transcription factors, and signaling pathways. These results demonstrate that DNA methylation is involved in the LT-regulated albino processes of ZH2. Changes in DNA methylation levels were associated with changes in gene expression levels, affecting the structure and function of chloroplasts, which may have a phenotypic impact on shoot and leaf color.

Differentially methylated genomic regions of lettuce seeds relate to divergence across morphologically distinct horticultural types.

Heritable cytosine methylation plays a role in shaping plant phenotypes; however, no information is available about DNA methylation in cultivated lettuce (Lactuca sativa), one of the most important leafy vegetables. Whole-genome bisulfite sequencing (WGBS) performed on seeds of 95 accessions from eight morphologically distinct horticultural types (Batavia, butterhead, iceberg, Latin, leaf, oilseed, romaine and stem) revealed a high level of methylation in lettuce genome with an average methylation of 90.6 % in the CG context, 72.9 % in the CHG context and 7.5 % in the CHH context. Although WGBS did not show substantial differences in overall methylation levels across eight horticultural types, 350 differentially methylated regions (DMR) were identified. Majority of the 41 pivotal DMR overlapped with genomic features predicted or confirmed to be involved in plant growth and development. These results provide the first insight into lettuce DNA methylation and indicate a potential role for heritable variation in cytosine methylation in lettuce morphology. The results reveal that differences in methylation profiles of morphologically distinct horticultural types are already detectable in seeds. Identified DMR can be a focus of the future functional studies.

Warmer temperature during asexual reproduction induce methylome, transcriptomic, and lasting phenotypic changes in Fragaria vesca ecotypes.

Plants must adapt with increasing speed to global warming to maintain their fitness. One rapid adaptation mechanism is epigenetic memory, which may provide organisms sufficient time to adapt to climate change. We studied how the perennial Fragaria vesca adapted to warmer temperatures (28°C vs. 18°C) over three asexual generations. Differences in flowering time, stolon number, and petiole length were induced by warmer temperature in one or more ecotypes after three asexual generations and persisted in a common garden environment. Induced methylome changes differed between the four ecotypes from Norway, Iceland, Italy, and Spain, but shared methylome responses were also identified. Most differentially methylated regions (DMRs) occurred in the CHG context, and most CHG and CHH DMRs were hypermethylated at the warmer temperature. In eight CHG DMR peaks, a highly similar methylation pattern could be observed between ecotypes. On average, 13% of the differentially methylated genes between ecotypes also showed a temperature-induced change in gene expression. We observed ecotype-specific methylation and expression patterns for genes related to gibberellin metabolism, flowering time, and epigenetic mechanisms. Furthermore, we observed a negative correlation with gene expression when repetitive elements were found near (±2kb) or inside genes. In conclusion, lasting phenotypic changes indicative of an epigenetic memory were induced by warmer temperature and were accompanied by changes in DNA methylation patterns. Both shared methylation patterns and transcriptome differences between F. vesca accessions were observed, indicating that DNA methylation may be involved in both general and ecotype-specific phenotypic variation.

Characteristics of Salvia miltiorrhiza methylome and the regulatory mechanism of DNA methylation in tanshinone biosynthesis.

Salvia miltiorrhiza is a model medicinal plant with significant economic and medicinal value. Its roots produce a group of diterpenoid lipophilic bioactive components, termed tanshinones. Biosynthesis and regulation of tanshinones has attracted widespread interest. However, the methylome of S. miltiorrhiza has not been analysed and the regulatory mechanism of DNA methylation in tanshinone production is largely unknown. Here we report single-base resolution DNA methylomes from roots and leaves. Comparative analysis revealed differential methylation patterns for CG, CHG, and CHH contexts and the association between DNA methylation and the expression of genes and small RNAs. Lowly methylated genes always had higher expression levels and 24-nucleotide sRNAs could be key players in the RdDM pathway in S. miltiorrhiza. DNA methylation variation analysis showed that CHH methylation contributed mostly to the difference. Go enrichment analysis showed that diterpenoid biosynthetic process was significantly enriched for genes with downstream overlapping with hypoCHHDMR in July_root when comparing with those in March_root. Tanshinone biosynthesis-related enzyme genes, such as DXS2, CMK, IDI1, HMGR2, DXR, MDS, CYP76AH1, 2OGD25, and CYP71D373, were less CHH methylated in gene promoters or downstream regions in roots collected in July than those collected in March. Consistently, gene expression was up-regulated in S. miltiorrhiza roots collected in July compared with March and the treatment of DNA methylation inhibitor 5-azacytidine significantly promoted tanshinone production. It suggests that DNA methylation plays a significant regulatory role in tanshinone biosynthesis in S. miltiorrhiza through changing the levels of CHH methylation in promoters or downstreams of key enzyme genes.

On the prediction of non-CG DNA methylation using machine learning.

DNA methylation can be detected and measured using sequencing instruments after sodium bisulfite conversion, but experiments can be expensive for large eukaryotic genomes. Sequencing nonuniformity and mapping biases can leave parts of the genome with low or no coverage, thus hampering the ability of obtaining DNA methylation levels for all cytosines. To address these limitations, several computational methods have been proposed that can predict DNA methylation from the DNA sequence around the cytosine or from the methylation level of nearby cytosines. However, most of these methods are entirely focused on CG methylation in humans and other mammals. In this work, we study, for the first time, the problem of predicting cytosine methylation for CG, CHG and CHH contexts on six plant species, either from the DNA primary sequence around the cytosine or from the methylation levels of neighboring cytosines. In this framework, we also study the cross-species prediction problem and the cross-context prediction problem (within the same species). Finally, we show that providing gene and repeat annotations allows existing classifiers to significantly improve their prediction accuracy. We introduce a new classifier called AMPS (annotation-based methylation prediction from sequence)that takes advantage of genomicannotations to achieve higher accuracy.

The promoters of OsGLP genes exhibited differentially methylated sites under drought and salt stress in rice cultivars

DNA methylation at cytosine residues governs the regulation of stress responsive genes in plant for self-protection against various environmental abiotic stresses. Here, we analyzed the epigenetic consequences of drought and salinity on cytosine methylation dynamics of promoter regions of stress responsive Germin-like protein genes in rice (Oryza sativa). The bisulfite sequencing technique was employed to identify differential methylation status at cytosine residues in selected promoter regions of three OsGLP genes (OsGLP4-1, OsGLP8-10 and OsGLP8-12) in leaves and roots of two elite Indica rice cultivars (tolerant KS282 and sensitive Super Basmati) under drought and salt stress. Our results identified cultivar, tissue and stress-dependent differentially methylated cytosine residues, however, the extent of methylation was found to be different depending upon CGN, CHG and CHH sequence contexts. Among all three OsGLP genes, the promoter region of OsGLP8-12 was detected with the most heavily methylated and significantly differential methylated sites depending upon types of variety, tissue or stress conditions. However, no methylated site was detected in the promoter region of OsGLP4-1. Moreover, in the promoter regions of OsGLP8-10 and OsGLP8-12, several differentially methylated sites in response to stress treatments were identified either near or within cis-regulatory elements (CREs) related to abiotic stress. This indicated the association between methylation in the promoter regions and regulation of OsGLP genes which might be a key mechanism associated with their regulation under abiotic stresses in contrasting rice cultivars.

DNA methylation trajectories during innate and adaptive immune responses of human B lymphocytes.

B lymphocytes can engage in either a rapid T cell-independent pathway (TI) or a delayed long-lasting T cell-independent (TD) response through highly ordered transcriptional programs, yet the detailed underlying mechanisms are unclear. Since DNA methylation plays a key role in controlling gene expression and lineage specification, we explored the dynamics of whole-genome cytosine modifications during the ex vivo response of human B cells isolated from normal individuals by negative selection. We found that B cell differentiation following TI and TD signalling is accompanied by extensive remodelling of the epigenome, including global gain and loss of DNA methylation. The epigenetic changes map to different regions of the B cell genome, including non- C-phosphate-G CpG islands, indicating that modifications of distal regulatory elements likely regulate specific gene transcription in B cells. Non-CpG methylation also occurs in differentiating human B cells, suggesting that this DNA modification is involved in transcriptional regulation of B cell genes with promoters exhibiting a low-density methylation, possibly by changing the chromatin shape that could have an impact on gene expression. Most strikingly, compared to TD activation, stimulation of B cells through an innate pathway induced higher levels of DNA methylation modifications at CpG, CHG and CGG contexts, supporting the view that DNA methylation modifications are used in distinct trajectories to specify the TI and TD B lymphocyte responses.

Epigenetic modification mechanisms of chloroplasts mutants in pineapple leaves during somatic regeneration

Somaclonal variation in tissue culture is a common phenomenon induced by various external or internal environmental conditions, resulting in heritable or non-heritable alterations in gene expression. One crucial mechanism involved in plant growth and development is epigenetic regulation. A highly dynamic epigenome can respond to environmental changes by regulating gene expression. DNA methylation is one of these epigenetic modifications that can alter gene expression in tissue-cultured pineapple plants. The underlying mechanism of such somaclonal variations in pineapple and the epigenetic regulation involvement in somaclonal variations has not been studied. This study performed DNA methylome and transcriptome sequencing of wild-type (WT) and mutant pineapple plants (WS, HW, and TW). We observed altered DNA methylation patterns in chlorophyll development in the mutants. Specifically, we noticed that the methylation levels in the CHG and CHH contexts were lower in the gene body regions compared to the upstream and downstream regions. We identified several thousand differentially methylated regions (DMRs) located at the gene body regions, some of which overlapped with the differentially expressed genes (DEGs). Functional enrichment analyses suggested that these genes were involved in chlorophyll metabolism. Thus, our results revealed that the transcriptional regulation of many chlorophyll metabolic essential genes could be regulated by DNA methylation caused by somaclonal variations and provided insights into epigenetic mechanisms underlying the regulation of chlorosis in pineapple plants.