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DNA methylation in the non-CG context is widespread in the plant kingdom and abundant in mammalian tissues such as the brain and pluripotent cells. Non-CG methylation in Arabidopsis thaliana is coordinately regulated by DOMAINS REARRANGED METHYLTRANSFERASE (DRM) and CHROMOMETHYLASE (CMT) proteins but has yet to be systematically studied in major crops due to difficulties in obtaining genetic materials. Here, utilizing the highly efficient multiplex CRISPR-Cas9 genome-editing system, we created single- and multiple-knockout mutants for all the nine DNA methyltransferases in rice (Oryza sativa) and profiled their whole-genome methylation status at single-nucleotide resolution. Surprisingly, the simultaneous loss of DRM2, CHROMOMETHYLASE3 (CMT2), and CMT3 functions, which completely erases all non-CG methylation in Arabidopsis, only partially reduced it in rice. The regions that remained heavily methylated in non-CG contexts in the rice Os-dcc (Osdrm2/cmt2/cmt3a) triple mutant had high GC contents. Furthermore, the residual non-CG methylation in the Os-dcc mutant was eliminated in the Os-ddccc (Osdrm2/drm3/cmt2/cmt3a/cmt3b) quintuple mutant but retained in the Os-ddcc (Osdrm2/drm3/cmt2/cmt3a) quadruple mutant, demonstrating that OsCMT3b maintains non-CG methylation in the absence of other major methyltransferases. Our results showed that OsCMT3b is subfunctionalized to accommodate a distinct cluster of non-CG-methylated sites at highly GC-rich regions in the rice genome., (© American Society of Plant Biologists 2021. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Allopolyploidization, which entails interspecific hybridization and whole genome duplication (WGD), is associated with emergent genetic and epigenetic instabilities that are thought to contribute to adaptation and evolution. One frequent genomic consequence of nascent allopolyploidization is homoeologous exchange (HE), which arises from compromised meiotic fidelity and generates genetically and phenotypically variable progenies. Here, we used a genetically tractable synthetic rice segmental allotetraploid system to interrogate genome-wide DNA methylation and gene expression responses and outcomes to the separate and combined effects of hybridization, WGD and HEs. Progenies of the tetraploid rice were genomically diverse due to genome-wide HEs that affected all chromosomes, yet they exhibited overall methylome stability. Nonetheless, regional variation of cytosine methylation states was widespread in the tetraploids. Transcriptome profiling revealed genome-wide alteration of gene expression, which at least in part associates with changes in DNA methylation. Intriguingly, changes of DNA methylation and gene expression could be decoupled from hybridity and sustained and amplified by HEs. Our results suggest that HEs, a prominent genetic consequence of nascent allopolyploidy, can exacerbate, diversify and perpetuate the effects of allopolyploidization on epigenetic and gene expression variation, and hence may contribute to allopolyploid evolution., (© 2019 The Authors. New Phytologist © 2019 New Phytologist Trust.)

Pleurotus tuoliensis (Pt) and P. eryngii var. eryngii (Pe) are important edible mushrooms. The epigenetic and gene expression signatures characterizing major developmental transitions in these two mushrooms remain largely unknown. Here, we report global analyses of DNA methylation and gene expression in both mushrooms across three major developmental transitions, from mycelium to primordium and to fruit body, by whole-genome bisulfite sequencing (WGBS) and RNA-seq-based transcriptome profiling. Our results revealed that in both Pt and Pe the landscapes of methylome are largely stable irrespective of genomic features, e.g., in both protein-coding genes and transposable elements (TEs), across the developmental transitions. The repressive impact of DNA methylation on expression of a small subset of genes is likely due to TE-associated effects rather than their own developmental dynamics. Global expression of gene orthologs was also broadly conserved between Pt and Pe, but discernible interspecific differences exist especially at the fruit body formation stage, and which are primarily due to differences in trans-acting factors. The methylome and transcriptome repertories we established for the two mushroom species may facilitate further studies of the epigenetic and transcriptional regulatory mechanisms underpinning gene during development in Pleurotus and related genera.

Rationale: The incidence of gestational diabetes mellitus (GDM) is increasing worldwide. However, whether and how GDM exposure induces fetal programming of adult cardiac dysfunctional phenotype, especially the underlying epigenetic molecular mechanisms and theranostics remain unclear. To address this problem, we developed a late GDM rat model. Methods: Pregnant rats were made diabetic on day 12 of gestation by streptozotocin (STZ). Experiments were conducted in 6 weeks old offspring. Results: There were significant increases in ischemia-induced cardiac infarction and gender-dependent left ventricular (LV) dysfunction in male offspring in GDM group as compared to controls. Exposure to GDM enhanced ROS level and caused a global DNA methylation in offspring cardiomyocytes. GDM attenuated cardiac Sirt 1 protein and p-Akt/Akt levels, but enhanced autophagy-related proteins expression (Atg 5 and LC3 II/LC3 I) as compared to controls. Ex-vivo treatment of DNA methylation inhibitor, 5-Aza directly inhibited Dnmt3A and enhanced Sirt 1 protein expression in fetal hearts. Furthermore, treatment with antioxidant, N-acetyl-cysteine (NAC) in offspring reversed GDM-mediated DNA hypermethylation, Sirt1 repression and autophagy-related gene protein overexpression in the hearts, and rescued GDM-induced deterioration in heart ischemic injury and LV dysfunction. Conclusion: Our data indicated that exposure to GDM induced offspring cardiac oxidative stress and DNA hypermethylation, resulting in an epigenetic down-regulation of Sirt1 gene and aberrant development of heart ischemia-sensitive phenotype, which suggests that Sirt 1-mediated signaling is the potential therapeutic target for the heart ischemic disease in offspring., Competing Interests: Competing Interests: The authors have declared that no competing interest exists.

Objective: Diabetes exerts adverse effects on the initiation or progression of diabetes and metabolic syndrome in the next generation. In past studies, limited attention has been given to the fathers' role in shaping the metabolic landscape of offspring. Our study was designed to investigate how paternal hyperglycemia exerts an intergenerational effect in mammals as well as the underlying mechanisms., Methods: Hyperglycemia was introduced in male rats by intraperitoneally injected streptozotocin and these males were bred with healthy females to generate offspring. The metabolic profiles of the progeny were assessed; DNA methylation profiles and gene expression were investigated. Mutagenesis constructs of the Ppara promoter region, and a luciferase reporter assay were used to determine transcription factor binding sites (TFBSs) and the effects of hypermethylation on Ppara transcription., Results: Paternal hyperglycemia induced increased liver weight, and plasma TC, TG, LDL, accumulation of triglycerides in the liver. We discovered that CpG 13 in the amplified promoter region (-852 to -601) of Ppara was hypermethylated in adult offspring liver and expression of Ppara, Acox1, Cpt-1α, and Cd36 was down regulated. Hypermethylation of CpG site 13 in the Ppara promoter inhibited the gene transcription, probably through abrogation of SP1 binding. The same epigenetic alteration was discovered in the fetus (E16.5) liver of hyperglycemic father's progeny., Conclusions: Paternal hyperglycemia may induce epigenetic modification of Ppara in offspring's liver, probably through interaction with SP1 binding, causing impaired lipid metabolism. Our investigation may have implications for the understanding of father-offspring interactions with the potential to account for metabolic syndromes., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Gene and genome duplication fosters genetic novelty, but redundant gene copies would undergo mutational decay unless preserved via selective or neutral forces. Molecular mechanisms mediating duplicate preservation remain incompletely understood. Several recent studies showed an association between DNA methylation and expression divergence of duplicated genes and suggested a role of epigenetic mechanism in duplicate retention. Here, we compare genome-wide gene-body CG methylation (BCGM) and duplicate gene expression between a rice mutant null for OsMet1-2(a major CG methytransferase in rice) and its isogenic wild-type. We demonstrate a causal link between BCGM divergence and expression difference of duplicate copies. Interestingly, the higher- and lower-expressing copies of duplicates as separate groups show broadly different responses with respect to direction of expression alteration upon loss of BCGM. A role for BCGM in conditioning expression divergence between copies of duplicates generally holds for duplicates generated by whole genome duplication (WGD) or by small-scale duplication processes. However, differences are evident among these categories, including a higher proportion of WGD duplicates manifesting expression alteration, and differential propensities to lose BCGM by the higher- and lower-expression copies in the mutant. Together, our results support the notion that differential epigenetic marking may facilitate long-term retention of duplicate genes.

Objective: To study the effect of cigarette smoke on chromatin configuration, DNA methylation and expression of related genes in mice oocytes., Methods: The clean grade ICR female mice were divided into 3 groups, control group, low dose group and high dose group. Animals were placed inside a perspex chamber( 18L) filled up with the smoke produced by 0, 1, 2 cigarettes, one hour a time, twice daily for 4weeks. Nuclear staining with Hoechst 33342, to observe the quality of oocyte and chromatin configuration. Analysis of the DNA methylation patterns by indirect immunofluorescence. The expressions of DNA methyltransferases( DNMT) 1, DNMT3a and DNMT3b mRNA were detected by quantitative real time PCR., Results: With the increase of smoke concentration, the diameter of oocytes was significantly decreased( P <0. 05), the percentage of non surrounded nucleolus( NSN) chromatin configuration was significantly increased( P < 0. 05). In high dose group, the level of DNA methylation wassignificantly lower than that of control group( P < 0. 05). The expression of DNMT1 decreased with the increase of smoke concentration, but the expression level of DNMT3b was significantly decreased( P < 0. 05)., Conclusion: Cigarette smoke exposure may alter oocyte chromatin configuration, decrease the expression level of DNMTs, resulted in oocytes DNA methylation decreased, so as to decrease the quality of oocytes.

Associations between 3D chromatin architectures and epigenetic modifications have been characterized in animals. However, any impact of DNA methylation on chromatin architecture in plants is understudied, which is confined to Arabidopsis thaliana. Because plant species differ in genome size, composition, and overall chromatin packing, it is unclear to what extent findings from A. thaliana hold in other species. Moreover, the incomplete chromatin architectural profiles and the low‐resolution high‐throughput chromosome conformation capture (Hi‐C) data from A. thaliana have hampered characterizing its subtle chromatin structures and their associations with DNA methylation. We constructed a high‐resolution Hi‐C interaction map for the null OsMET1‐2 (the major CG methyltransferase in rice) mutant (osmet1‐2) and isogenic wild‐type rice (WT). Chromatin structural changes occurred in osmet1‐2, including intra‐/inter‐chromosomal interactions, compartment transition, and topologically associated domains (TAD) variations. Our findings provide novel insights into the potential function of DNA methylation in TAD formation in rice and confirmed DNA methylation plays similar essential roles in chromatin packing in A. thaliana and rice. [ABSTRACT FROM AUTHOR]

Summary: CG methylation (mCG) is essential for preserving genome stability in mammals, but this link remains obscure in plants. OsMET1‐2, a major rice DNA methyltransferase, plays critical roles in maintaining mCG in rice. Null mutation of OsMET1‐2 causes massive CG hypomethylation, rendering the mutant suitable to address the role of mCG in maintaining genome integrity in plants. Here, we analyzed mCG dynamics and genome stability in tissue cultures of OsMET1‐2 homozygous (−/−) and heterozygous (+/−) mutants, and isogenic wild‐type (WT). We found mCG levels in cultures of −/− were substantially lower than in those of WT and +/−, as expected. Unexpectedly, mCG levels in 1‐ and 3‐year cultures of −/− were 77.6% and 48.7% higher, respectively, than in shoot, from which the cultures were initiated, suggesting substantial regain of mCG in −/− cultures, which contrasts to the general trend of mCG loss in all WT plant tissue cultures hitherto studied. Transpositional burst of diverse transposable elements (TEs) occurred only in −/− cultures, although no elevation of genome‐wide mutation rate in the form of single nucleotide polymorphisms was detected. Altogether, our results establish an essential role of mCG in retaining TE immobility and hence genome stability in rice and likely in plants in general. Significance Statement: This study establishes a system in rice with diverse transposable elements (TEs) being mobile in real‐time. This, together with the associated genome, methylome and transcriptome data, may provide a valuable entry‐point for TE biology studies in plants. [ABSTRACT FROM AUTHOR]

Mercury (Hg) is an important hazardous pollutant that can cause phytotoxicity and harm human health through the food chain. Recently, rice (Oryza sativa L.) has been confirmed as a potential Hg bioaccumulator. Although the genetic and molecular mechanisms involved in heavy metal absorption and translocation in rice have been investigated for several heavy metals, Hg is largely neglected. Here, we analyzed one Hg-resistant line in rice (RHg) derived from a DNA methyltransferase-coding gene, OsMET1–2 heterozygous mutant. Compared with its isogenic wild-type (WT), RHg exhibited a significantly higher survival rate after Hg treatment, ameliorated oxidative damage, and lower Hg uptake and translocation. RNAseq-based comparative transcriptomic analysis identified 34 potential Hg resistance-related genes involved in phytohormone signaling, abiotic stress response, and zinc (Zn) transport. Importantly, the elevated expression of Hg resistance-related genes in RHg was highly correlated with DNA hypomethylation in their putative promoter regions. An ionomic analysis unraveled a negative correlation between Zn and Hg in roots. Moreover, Hg concentration was effectively decreased by exogenous application of Zn in Hg-stressed rice plants. Our findings indicate an epigenetic basis of Hg resistance and reveal an antagonistic relationship between Hg and Zn, providing new hints towards Hg detoxification in plants. Mercury (Hg) as an important hazardous pollutant adversely impacts the environment and jeopardizes human health, due to its chronicity, transferability, persistence, bioaccumulation and toxicity. In this paper, we identified 34 potential genes that may significantly contribute to Hg resistance in rice. We find the expression of Hg resistance-related genes was highly correlated with DNA hypomethylation in their putative promoter regions. Our results also revealed an antagonistic relationship between Hg and Zinc (Zn), providing new hints towards Hg detoxification in plants. Together, findings of this study extend our current understanding of Hg tolerance in rice and are informative to breed seed non–accumulating rice cultivars. [Display omitted] • Identified one Hg-resistant line (RHg) in rice derived from OsMET1-2 +/- . • RHg accumulated less ROS and Hg2+ compared with its isogenic wild-type. • Identified 34 potential Hg resistance-related genes in rice. • Hg induces DNA hypomethylation at promoter regions of upregulated candidate genes. • Revealed an antagonistic relationship between Hg and Zn. [ABSTRACT FROM AUTHOR]