H3K27me3 Levels Research Articles

EPO modified MSCs protects SH-SY5Y cells against ischemia/hypoxia-induced apoptosis via REST-dependent epigenetic remodeling

Hypoxic-ischemic encephalopathy (HIE) is a diffuse brain tissue injury caused by acute ischemia and hypoxia, and it is most commonly found in newborn infants but can also occur in adults. Mesenchymal stem cell (MSC) therapies have showed improved outcomes for treating HIE-induced neuronal defects. However, many key issues associated with poor cell viability and tolerance of grafted MSCs after HIE remain to be resolved. Genetic engineering could endow MSCs with more robust regenerative capacities. Our research, along with that of other scientists, has found that the expression of intracellular erythropoietin (EPO) in human umbilical cord MSCs (hUC-MSCs) increases proportionally with the duration of hypoxia exposure. Furthermore, we observed that EPO, when introduced into the EPO gene-modified hUC-MSCs, can be secreted into the extracellular space. However, the underlying mechanisms that support the neuroprotective effects of EPO-MSCs remain unclear. EPO-MSCs, hUC-MSCs, and NC-MSCs were identified by flow cytometry, osteogenic, and adipogenic differentiation assays. The oxygen-glucose deprivation (OGD)-induced SH-SY5Y cell-line was established, and five groups were set up: control, 24-h ischemia-hypoxia, co-cultured with hUC-MSCs, NC-MSCs, and EPO-MSCs after hypoxia. LEGENDplex™ multi-factor flow cytometry was used to detect the secretion of inflammatory factors in cell supernatants and cerebrospinal fluid. Chromosome-targeted excision and tagging (CUT&Tag) sequencing was applied to detect genomic H3K4me2 modifications, and conjoint analysis with transcriptome sequencing (RNA-seq) was performed. Lentiviral vector infection was used to construct SH-SY5Y cells with stable knockdown of RE1-silencing transcription factor (REST), and flow cytometry was used to detect alterations in apoptosis. Finally, the molecular mechanism underlying the neuroprotective and anti-apoptotic effects of EPO-MSCs was investigated using RNA sequencing, qRT-PCR, and western blot assays. Our results suggest that EPO-MSCs are genetically engineered to secrete significantly more EPO. EPO-MSCs treatment has anti-apoptotic properties and offers neuronal protection during ischemic-hypoxic injury. Furthermore, RNA-seq results suggest that multiple inflammation-related genes were down-regulated after EPO-MSCs treatment. Application of RNA-seq and CUT&Tag combined analysis found that the expressions of REST were significantly up-regulated. Lentiviral vector infection to construct REST knockdown SH-SY5Y failed to rescue apoptosis after hypoxia and co-culture with EPO-MSCs, and SETD2-mediated H3K36me3 protein level expression was reduced. EPO-MSCs may promote neuronal survival by affecting H3K4me2 and thus activating the expression of REST and TET3. EPO-MSCs also upregulated the modification level of SETD2-mediated H3K36me3 and regulated the expression of inflammation-related genes such as PLCG2, as well as apoptosis genes BCL2A1. To investigate the neuroprotective effects of EPO-modified hUC-MSCs and the underlying epigenetic regulatory mechanisms, this study aims to provide a theoretical foundation for the potential application of EPO gene-modified hUC-MSCs in the treatment of HIE.

Enriched H3K27Me3 on BMP4 suppresses the osteoblastic differentiation potential of BMSCs in diabetes mellitus

Diabetes mellitus has been widely acknowledged to have a negative effect on the osteoblastic differentiation of bone marrow-derived mesenchymal stem cells (BMSCs). However, the underlying epigenetic mechanisms associated with this process remain to be elucidated. The goal of the present study was to investigate the effect of diabetes mellitus on the osteoblastic differentiation of BMSCs and assess the role of histone methylation in the observed phenomena. The osteoblastic differentiation ability of BMSCs was shown to be decreased in diabetes mellitus, as indicated by alkaline phosphatase activity and the mRNA levels of osteoblast-related genes. Furthermore, diabetes mellitus caused an increased expression of the histone methylase EZH2 and the levels of H3K27Me3 and decreased the expression of the histone demethylase KDM6B, as demonstrated by qRT-PCR and western blotting. Furthermore, immunofluorescence staining suggested that both EZH2 and H3K27Me3 were primarily localized in the nucleus. In addition, chromatin immunoprecipitation assays indicated an increased presence of H3K27Me3 on the promoter region of the BMP4 gene. In summary, in the present study, we demonstrated that the osteoblastic differentiation of BMSCs is dramatically reduced in diabetes mellitus. In addition, upregulation of EZH2 expression and downregulation of KDM6B expression may not be enough to eliminate transcriptional repression mediated by H3K27Me3 on the promoter region of the BMP4 gene during the osteoblastic differentiation of BMSCs in diabetes mellitus.

KDM4B Histone Demethylase Inhibition Attenuates Tumorigenicity of Malignant Melanoma Cells by Overriding the p53-Mediated Tumor Suppressor Pathway.

Despite significant advances in the treatment of cutaneous melanoma (hereaftermelanoma), the prognosis remains less favorable due to therapeutic resistance, which is presumably linked to epigenetic dysregulation. We hypothesized that the histone lysine demethylase KDM4B could play a pivotal role in controlling therapy-resistant melanoma. To validate our hypothesis, we retrieved RNA sequencing data from the Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA)program and observed upregulation of KDM4B in both primary and metastatic melanoma, which was associated with poor survival. To explore its role, we used murine B16, human SK-MEL-5, and G-361 melanoma cells as in vitro models of melanoma. We found that KDM4B inhibition using NCGC00244536 increased global levels of H3K9me3 and downregulated the expressions of cell cycle progression-related genes Cdk1, Cdk4, Ccnb1, and Ccnd1. Moreover, genetic ablation of KDM4B or its chemical inhibition using NCGC00244536 reduced p53 production by upregulating MDM2, which enhances the proteolytic degradation of p53. Interestingly, despite the reduction of p53, these interventions augmented apoptosis and senescence-induced cell death by activating pathways downstream of p53, as evidenced by reduced levels of pro-survival Bcl-2 and Bcl-xL proteins and increased production of pro-apoptotic cleaved caspase-3, caspase-7, Bax, and the senescence inducer Cdkn1a. Compared to the FDA-approved anti-melanoma agent dacarbazine, NCGC00244536 exhibited more pronounced cytotoxic and antiproliferative effects in melanoma cells. Importantly, NCGC00244536 demonstrated minimal cytotoxicity to low Kdm4b-expressing mouse embryonic fibroblasts. In conclusion, our findings suggest that KDM4B inhibition can override the antitumor effect of p53, and potentially serve as a therapeutic strategy for melanoma.

LINE1 and PRC2 control nucleolar organization and repression of the 8C state in human ESCs.

The mechanisms that ensure developmental progression in the early human embryo remain largely unknown. Here, we show that the family of long interspersed nuclear element 1 (LINE1) transposons prevents the reversion of naive human embryonic stem cells (hESCs) to 8-cell-like cells (8CLCs). LINE1 RNA contributes to maintenance of H3K27me3 levels, particularly at chromosome 19 (Chr19). Chr19 is enriched for key 8C regulators, H3K27me3, and genes derepressed upon LINE1 knockdown or PRC2 inhibition. Moreover, Chr19 is strongly associated with the nucleolus in hESCs but less in 8CLCs. Direct inhibition of PRC2 activity induces the 8C program and leads to a relocalization of Chr19 away from the nucleolus. LINE1 KD or PRC2 inhibition induces nucleolar stress, and disruption of nucleolar architecture is sufficient to de-repress the 8C program. These results indicate that LINE1 RNA and PRC2 maintain H3K27me3-mediated gene repression and 3D nuclear organization to prevent developmental reversion of hESCs.

The expression and role of SUZ12 in lung adenocarcinoma.

SUZ12 is one of the core members of the polycomb repressive complex 2 (PRC2), but its expression and role in lung adenocarcinoma (LUAD) are unclear. We aimed to explore the expression, prognosis, biological functions and roles of SUZ12 in LUAD. The expression of SUZ12 was detected by immunohistochemical staining, qRT-PCR, and western blotting in LUAD tissues and cells. The biological functions and molecular mechanisms of SUZ12 were characterized by a range of invitro and invivo experiments. SUZ12 was overexpressed in LUAD tissues, and high SUZ12 expression was correlated with worse clinicopathological features and a poorer prognosis. Knockdown of SUZ12 significantly inhibited cell growth, colony formation, invasion, and migration, and induced apoptosis and G1/S phase arrest, while overexpression of SUZ12 had the opposite effects. Knockdown of SUZ12 decreased the tumorigenic capacity of A549 cells invivo. The expression of key signaling molecules related to the cell cycle, apoptosis, migration, and immunity were altered by the knockdown or overexpression of SUZ12. SUZ12 can directly bind to the Bax promoter region, EZH2 and H3K27me3 levels dependents on SUZ12. The expression levels of SUZ12 and Bax were negatively correlated in LUAD tissues. SUZ12 is a new oncogene related to the poor prognosis of LUAD. SUZ12 regulates LUAD progression by regulating the expression of related signaling molecules, and as a part of the PRC2 complex, it may bind to the Bax promoter to silence Bax expression.

Kdm4d mutant mice show impaired sperm motility and subfertility.

Regulation of gene expression through histone modifications underlies cell homeostasis and differentiation. Kdm4d and Kdm4dl exhibit a high degree of similarity and demethylate H3K9me3. However, the physiological functions of these proteins remain unclear. In this study, we generated Kdm4dl mutant mice and found that Kdm4dl was dispensable for mouse development. However, through the generation of Kdm4d mutant mice, we unexpectedly found that Kdm4d mutant male mice were subfertile because of impaired sperm motility. The absence of Kdm4d was associated with an altered distribution of H3K9me3 in round spermatids, suggesting that the Kdm4d-mediated adjustment of H3K9me3 levels is required to generate motile sperm. Further analysis revealed that the absence of Kdm4d did not affect the functionality of sperm nuclei in generating offspring. As KDM4D is specifically expressed in the human testes, our results suggest that changes in KDM4D expression or its activity may be a risk factor for human infertility.

Functional characterisation of BnaA02.TOP1α and BnaC02.TOP1α involved in true leaf biomass accumulation in Brassica napus L.

Leaves, as primary photosynthetic organs essential for high crop yield and quality, have attracted significant attention. The functions of DNA topoisomerase 1α (TOP1α) in various biological processes, including leaf development, in Brassica napus remain unknown. Here, four paralogs of BnaTOP1α, namely BnaA01.TOP1α, BnaA02.TOP1α, BnaC01.TOP1α and BnaC02.TOP1α, were identified and cloned in the B. napus inbred line 'K407'. Expression pattern analysis revealed that BnaA02.TOP1α and BnaC02.TOP1α, but not BnaA01.TOP1α and BnaC01.TOP1α, were persistently and highly expressed in B. napus true leaves. Preliminary analysis in Arabidopsis thaliana revealed that BnaA02.TOP1α and BnaC02.TOP1α paralogs, but not BnaA01.TOP1α and BnaC01.TOP1α, performed biological functions. Targeted mutations of four BnaTOP1α paralogs in B. napus using the CRISPR-Cas9 system revealed that BnaA02.TOP1α and BnaC02.TOP1α served as functional paralogs and redundantly promoted true leaf number and size, thereby promoting true leaf biomass accumulation. Moreover, BnaA02.TOP1α modulated the levels of endogenous gibberellins, cytokinins and auxins by indirectly regulating several genes related to their metabolism processes. BnaA02.TOP1α directly activated BnaA03.CCS52A2 and BnaC09.AN3 by facilitating the recruitment of RNA polymerase II and modulating H3K27me3, H3K36me2 and H3K36me3 levels at these loci and indirectly activated the BnaA08.PARL1 expression, thereby positively controlling the true leaf size in B. napus. Additionally, BnaA02.TOP1α indirectly activated the BnaA07.PIN1 expression to positively regulate the true leaf number. These results reveal the important functions of BnaTOP1α and provide insights into the regulatory network controlling true leaf biomass accumulation in B. napus.

Epigenetic mechanism of EZH2-mediated histone methylation modification in regulating ferroptosis of alveolar epithelial cells in sepsis-induced acute lung injury.

Sepsis-induced acute lung injury (SI-ALI) leads to significant deaths in critically ill patients worldwide. This study explores the mechanism of EZH2 regulating ferroptosis of alveolar epithelial cells (AECs) in SI-ALI. In vitro cell model and in vivo mouse lung injury model of sepsis were established. EZH2 expression in lung tissues was intervened by sh-EZH2, followed by H&E staining observation of lung tissue pathological changes. EZH2, H3K27me3, USP10, GPX4, and ACSL4 expressions were determined by qRT-PCR or Western blot. ROS, GSH, and iron ion levels were detected using fluorescent labeling and reagent kits, respectively. ChIP analyzed the enrichment of EZH2 and H3K27me3 on USP10 promoter. The binding between USP10 and GPX4, and the ubiquitination level of GPX4 were detected using Co-IP. EZH2 was highly expressed in lung tissues of SI-ALI mice. EZH2 silencing alleviated ALI and ferroptosis of AECs; EZH2 increased the H3K27me3 level on USP10 promoter through histone methylation. USP10 stabilized GPX4 protein expression through ubiquitination; inhibition of USP10 partially reversed the inhibitory effect of EZH2 silencing on ferroptosis of AECs. In conclusion, EZH2 depresses USP10 expression by promoting histone H3K27me3 modification on USP10 promoter, thereby enhancing ubiquitination degradation of GPX4 and ultimately facilitating ferroptosis of AECs in sepsis.

Characterisation and evolution of the PRC2 complex and its functional analysis under various stress conditions in rice

The polycomb repressive complex 2 (PRC2) is a chromatin-associated methyltransferase responsible for catalysing the trimethylation of H3K27, an inhibitory chromatin marker associated with gene silencing. This enzymatic activity is crucial for normal organismal development and the maintenance of gene expression patterns that preserve cellular identity, subsequently influencing plant growth and abiotic stress responses. Therefore, in this study, we investigated the evolutionary characteristics and functional roles of PRC2 in plants. We identified 209 PRC2 genes, including E(z), Su(z), Esc, and Nurf55 families, using 18 representative plant species and revealed that recent gene replication events have led to an expansion in the Nurf55 family, resulting in a greater number of members compared to the E(z), Su(z), and Esc families. Furthermore, protein structure and motif composition analyses highlighted the potential functional site regions within PRC2 members. In addition, we selected rice, a representative monocotyledonous plant, as the model species for food crops. Our findings revealed that SDG711, SDG718, and MSI1-5 genes were induced by abscisic acid (ABA) and/or methyl jasmonate (MeJA) hormones, suggesting that these genes play an important role in abiotic stress and disease resistance. Further experiments involving rice blast fungus treatments confirmed that the expression of SDG711 and MSI1-5 was induced by Magnaporthe oryzae strain GUY11. Multiple protein interaction assays revealed that the M. oryzae effector AvrPiz-t interacts with PRC2 core member SDG711 to increase H3K27me3 levels. Notably, inhibition of PRC2 or mutation of SDG711 enhanced rice resistance to M. oryzae. Collectively, these results provide new insights into PRC2 evolution in plants and its significant functions in rice.

H3K4me3 changes occur in cell wall genes during the development of Fagopyrum tataricum morphogenic and non-morphogenic calli.

Epigenetic changes accompany the dynamic changes in the cell wall composition during the development of callus cells. H3K4me3 is responsible for active gene expression and reaction to environmental cues. Chromatin immunoprecipitation (ChIP) is a powerful technique for studying the interplay between epigenetic modifications and the DNA regions of interest. In combination with sequencing, it can provide the genome-wide enrichment of the specific epigenetic mark, providing vital information on its involvement in the plethora of cellular processes. Here, we describe the genome-wide distribution of H3K4me3 in morphogenic and non-morphogenic callus of Fagopyrum tataricum. Levels of H3K4me3 were higher around the transcription start site, in agreement with the role of this mark in transcriptional activation. The global levels of methylation were higher in the non-morphogenic callus, which indicated increased gene activation compared to the morphogenic callus. We also employed ChIP to analyse the changes in the enrichment of this epigenetic mark on the cell wall-related genes in both calli types during the course of the passage. Enrichment of H3K4me3 on cell wall genes was specific for callus type, suggesting that the role of this mark in cell-wall remodelling is complex and involved in many processes related to dedifferentiation and redifferentiation. This intricacy of the cell wall composition was supported by the immunohistochemical analysis of the cell wall epitopes' distribution of pectins and extensins. Together, these data give a novel insight into the involvement of H3K4me3 in the regeneration processes in F. tataricum in vitro callus tissue culture.

The role of trimethylation on histone H3 lysine 27 (H3K27me3) in temozolomide resistance of glioma

Temozolomide (TMZ) is the first-line chemotherapeutic agent for malignant glioma, but its resistance limited the benefits of the treated patients. In this study, the role and significance of trimethylation of histone H3 lysine 27 (H3K27me3) in TMZ resistance were investigated. Data from twenty advanced glioma patients were collected, and their pathological samples were analyzed for H3K27me3 levels. TMZ sensitivity was compared between glioma cells U87 and TMZ-resistant cells U87TR, with H3K27me3 levels determined in both cells. The effects of H3K27me3 demethylases inhibitor GSK-J4, combined with TMZ, were assessed on the proliferation and migration of U87TR cells. The results indicated that a high level of H3K27me3 predicts longer disease free survival (DFS) and overall survival (OS) in glioma patients receiving TMZ treatment. The H3K27me3 level was lower in U87TR cells compared to U87 cells. GSK-J4 increased the H3K27me3 level in U87TR cells and decreased their resistance to TMZ. In summary, this study identified a novel marker of TMZ resistance in glioma and provided a new strategy to address this challenge. These findings are significant for improving the clinical treatment of glioma in the future.

Eed-dependent histone modification orchestrates the iNKT cell developmental program alleviating liver injury.

Polycomb repressive complex 2 (PRC2) is an evolutionarily conserved epigenetic modifier responsible for tri-methylation of lysine 27 on histone H3 (H3K27me3). Previous studies have linked PRC2 to invariant natural killer T (iNKT) cell development, but its physiological and precise role remained unclear. To address this, we conditionally deleted Eed, a core subunit of PRC2, in mouse T cells. The results showed that Eed-deficient mice exhibited a severe reduction in iNKT cell numbers, particularly NKT1 and NKT17 cells, while conventional T cells and NKT2 cells remained intact. Deletion of Eed disrupted iNKT cell differentiation, leading to increased cell death, which was accompanied by a severe reduction in H3K27me3 levels and abnormal expression of Zbtb16, Cdkn2a, and Cdkn1a. Interestingly, Eed-deficient mice were highly susceptible to acetaminophen-induced liver injury and inflammation in an iNKT cell-dependent manner, highlighting the critical role of Eed-mediated H3K27me3 marks in liver-resident iNKT cells. These findings provide further insight into the epigenetic orchestration of iNKT cell-specific transcriptional programs.

Carbohydrate response element-binding protein (ChREBP) mediates decreased SNAP25 expression in islets from diabetic Goto-Kakizaki (GK) rats.

SNAP25 plays an essential role in the glucose-stimulated insulin secretion (GSIS) of pancreatic β-cells. Carbohydrate response element-binding protein (ChREBP) is an important transcription factor in β-cells and, in this study, we aimed to explore whether ChREBP regulates SNAP25 expression in β-cells. We show that diabetic Goto-Kakizaki (GK) rats exhibited impaired insulin secretion and hyperglycemia, along with decreased SNAP25 expression and ChREBP phosphorylation in islets. SNAP25 knockdown decreased GSIS in β-cells, while SNAP25 overexpression increased GSIS in β-cells. Activation or overexpression of ChREBP led to reduced SNAP25 expression and subsequent suppression of GSIS. Conversely, ChREBP knockdown mitigated the reduction in SNAP25 expression caused by high glucose. Mechanistically, the activation of ChREBP by high glucose increased its occupancy and decreased the level of H3K4me3 at the Snap25 promoter. Our findings reveal the novel regulatory mechanisms of SNAP25 expression in β-cells and suggest that SNAP25 may be involved in the regulation of β-cell secretory function controlled by ChREBP.

Abstract B082: Oncogenic KRAS-mediated epigenetic reprogramming is altered by loss of Activating Transcription Factor 3

Abstract Introduction: Over 90% of PDAC patients harbor an activating KRAS mutation, the most common form being KRASG12D. However, additional genetic mutations and environmental events, including chronic or hereditary pancreatitis, are needed for KRAS mutations to lead to PDAC. Our laboratory showed Activating Transcription Factor 3 (ATF3) was required for repressing genes that stabilize the mature acinar cell phenotype and for KRASG12D-driven progression to advanced PanIN lesions. However, ATF3 contributes to gene activation and repression and has been linked to numerous transcription factors and epigenetic mediators, and how ATF3 affects PDAC progression remains unknown. We hypothesize that KRASG12D promotes ATF3-driven histone acetylation in precursor PanINs lesions. Methods: C57Bl/6l mice allowing for acinar-inducible KRASG12D combined with (Ptf1acreERTKRASG12D) or without (Ptf1acreERTKRASG12DAtf3-/-; APK) Atf3 deletion were treated with tamoxifen to induce KRASG12D. After 10 days, mice were treated with cerulein to induce injury and PanIN progression. Mice were sacrificed after two weeks and 3D organoid lines developed. RNA-seq and chromatin immunoprecipitation for H3K27 acetylation (H3K27ac) followed by sequencing (ChIP-seq) was performed. Bioinformatic analysis was used to identify differentially enriched pathways. Levels of acetyl-CoA, the substrate for acetylation, was compared between lines along with levels of H3K27ac, H3K4me1, H3K4me3, and H3K9me3 through western blots. Results and Discussion: KRASG12D expression promoted epigenetic reprogramming in PanINs, including dysregulation of H3K27ac enrichment. ChIP-seq results showed the absence of ATF3 reduces H3K27 acetylation preferentially at gene promoters and pathway analysis showed differential enrichment and activation of KRAS signaling. Mechanistically, ATF3-deleted mice have lower acetyl-CoA levels, linking ATF3 to metabolic pathways as a possible mechanism for gene regulation. Comparing genes with an ATF3-dependent acetylation pattern to a curated list of KRAS-targeted genes identified SMARCC2 (BAF170), a SWI/SNF complex subunit, and KDM1B, a histone demethylase affecting H3K4, suggesting a potential role for ATF3 in widespread epigenetic regulation. Public sequencing data from PDAC patients, supports these findings as ATF3 expression is positively correlated with expression of several SWI/SNF complex subunits and demethylases. Conclusion: This study highlights a role for ATF3- mediated epigenetic dysregulatio. In oncogenic KRAS-dependent PDAC. Targeting this ATF3- mediated epigenetic landscape could be a promising new therapeutic avenue. Citation Format: Fatemeh Mousavi, Christopher L Pin, Parisa Shooshtari. Oncogenic KRAS-mediated epigenetic reprogramming is altered by loss of Activating Transcription Factor 3 [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B082.

Age-related histone H3.3 accumulation associates with a repressive chromatin in mouse tibialis anterior muscle

The present study aimed to investigate age-related changes in histone variant H3.3 and its role in the aging process of mouse tibialis anterior muscle. H3.3 level significantly increased with age and correlated with H3K27me3 level. Acute exercise successfully upregulated the target gene expression in 8-wk-old mice, whereas no upregulation was noted in 53-wk-old mice. H3K27me3 level was increased at these loci in response to acute exercise in 8-wk-old mice. However, in 53-wk-old mice, H3.3 and H3K27me3 levels were increased at rest and were not affected by acute exercise. Furthermore, forced H3.3 expression in the skeletal muscle of 8-wk-old mice led to a gradual improvement in motor function. The results suggest that age-related H3.3 accumulation induces the formation of repressive chromatin in the mouse tibialis anterior muscle. However, H3.3 accumulation also appears to play a positive role in enhancing skeletal muscle function.

Mechanism of KMT2D-mediated epigenetic modification in IL-1β-induced nucleus pulposus cell degeneration.

Intervertebral disc (IVD) degeneration (IVDD) is characterized by structural destruction accompanied by accelerated signs of aging. This study aimed to investigate the mechanism of lysine methyltransferase 2D (KMT2D) in the proliferation, apoptosis, and inflammation of nucleus pulposus cells (NPCs) in IVDD. Mouse-derived NPCs were cultured and induced with interleukin-1 beta (IL-1β) to establish cell models. KMT2D expression was detected by western blot and reverse transcription-quantitative polymerase chain reaction (RT-qPCR). KMT2D expression was interfered with, and the contents of IL-18, IL-6, and tumor necrosis factor (TNF) were detected by enzyme-linked immunosorbent assay. Cell proliferation, apoptosis, and the expression of miR-133a-5p and 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 2 (PFKFB2) were measured. The enrichment of KMT2D and Histone 3 Lysine 4 monomethylation/dimethylation (H3K4me1/2) on the miR-133a-5p promoter was analyzed by chromatin immunoprecipitation and qPCR. The binding of miR-133a-5p and PFKFB2 was analyzed by a dual-luciferase assay. IL-1β treatment promoted KMT2D expression in NPCs. KMT2D knockdown reduced inflammation and apoptosis and promoted the proliferation of IL-1β-induced NPCs. Mechanistically, KMT2D upregulated miR-133a-5p expression by increasing the level of H3K4me2 at the miR-133a-5p promoter, thereby promoting the binding between miR-133a-5p and PFKFB2 and downregulating the transcription of PFKFB2. miR-133a-5p overexpression or PFKFB2 knockdown alleviated the protective effect of KMT2D knockdown on IL-1β-induced NPCs. KMT2D promoted miR-133a-5p expression through H3K4me2 methylation, thereby promoting the binding of miR-133a-5p to PFKFB2, reducing the mRNA level of PFKFB2, promoting inflammation and apoptosis of IL-1β-induced NPCs, and inhibiting NPC proliferation.

Epigenetic alteration of uterine Leukemia Inhibitory Factor gene after glyphosate or a glyphosate-based herbicide exposure in rats

Glyphosate-based herbicides (GBHs) or its active ingredient, glyphosate (Gly), induce implantation failure in rats. We aimed to elucidate a mechanism of action of these compounds assessing the transcriptional and epigenetic status of the receptivity marker, leukemia inhibitory factor (Lif) gene. F0 rats were orally exposed to GBH or Gly at 3.8 or 3.9 mg Gly/kg/day, respectively, from gestational day (GD) 9 until weaning. F1 females were mated and uterine samples collected at GD5. Methylation-sensitive restriction enzymes (MSRE) sites and transcription factors were in silico predicted in regulatory regions of Lif gene. DNA methylation status and histone modifications (histone 3 and 4 acetylation (H3Ac and H4Ac) and H3 lysine-27-trimethylation (H3K27me3)) were assessed. GBH and Gly decreased Lif mRNA levels and caused DNA hypermethylation. GBH increased H3Ac levels, whereas Gly reduced them; both compounds enhanced H3K27me3 levels. Finally, both GBH and Gly induced similar epigenetic alterations in the regulatory regions of Lif.

The SET-Domain-Containing Protein MdSDG26 Negatively Regulates Alternaria alternata Resistance in Apple.

Apple leaf spot is one of the most devastating diseases in the apple industry, caused by Alternaria alternata f. sp mali (A. alternata). SET-domain group (SDG) proteins function as the histone methyltransferases and participate in plant development and stress responses. However, whether SDG proteins are associated with A. alternata resistance is largely unclear. Here, we describe the pathogen-inducible MdSDG26 gene in apple (Malus × domestica). MdSDG26 has two transcript variants that function similarly in catalyzing histone methylation and A. alternata resistance. Transient overexpression of MdSDG26 increased the global levels of H3K4me3 and H3K36me3, whereas knockdown of MdSDG26 only reduced the H3K36me3 level. Transcriptome analysis revealed that MdSDG26 affected the genome-wide transcriptome changes in response to A. alternata infection. ChIP-qPCR analysis demonstrated that MdSDG26 modulates the levels of H3K36me3 and H3K4me3 at both the promoter and exon regions of MdNTL9. As a negative regulator of A. alternata resistance in apples, MdNTL9 plays a pivotal role in MdSDG26-mediated resistance to A. alternata. Therefore, our findings provide compelling evidence for the regulatory function of MdSDG26 in histone methylation and its molecular role in conferring resistance to A. alternata.

MAT2A is essential for zygotic genome activation by maintaining of histone methylation in porcine embryos

Methionine adenosyltransferase 2A (MAT2A) is an essential enzyme in the methionine cycle that generates S-adenosylmethionine (SAM) by reacting with methionine and ATP. SAM acts as a methyl donors for histone and DNA methylation, which plays key roles in zygotic genome activation (ZGA). However, the effects of MAT2A on porcine ZGA remain unclear. To investigate the function of MAT2A and its underlying mechanism in porcine ZGA, MAT2A was knocked down by double-stranded RNA injection at the 1-cell stage. MAT2A is highly expressed at every stage of porcine embryo development. The percentages of four-cell-stage embryos and blastocysts were lower in the MAT2A-knockdown (KD) group than in the control group. Notably, depletion of MAT2A decreased the levels of H3K4me2, H3K9me2/3, and H3K27me3 at the four-cell stage, whereas MAT2A KD reduced the transcriptional activity of ZGA genes. MAT2A KD decreased embryonic ectoderm development (EED) and enhancer of zeste homolog 2 (EZH2) expression. Exogenous SAM supplementation rescued histone methylation levels and developmental arrest induced by MAT2A KD. Additionally, MAT2A KD significantly increased DNA damage and apoptosis. In conclusion, MAT2A is involved in regulating transcriptional activity and is essential for regulating histone methylation during porcine ZGA.

DNA methylation and histone modifications associated with antipsychotic treatment: a systematic review.

Antipsychotic medications are essential when treating schizophrenia spectrum and other psychotic disorders, but the efficacy and tolerability of these medications vary from person to person. This interindividual variation is likely mediated, at least in part, by epigenomic processes that have yet to be fully elucidated. Herein, we systematically identified and evaluated 65 studies that examine the influence of antipsychotic drugs on epigenomic changes, including global methylation (9 studies), genome-wide methylation (22 studies), candidate gene methylation (16 studies), and histone modification (18 studies). Our evaluation revealed that haloperidol was consistently associated with increased global hypermethylation, which corroborates with genome-wide analyses, mostly performed by methylation arrays. In contrast, clozapine seems to promote hypomethylation across the epigenome. Candidate-gene methylation studies reveal varying effects post-antipsychotic therapy. Some genes like Glra1 and Drd2 are frequently found to undergo hypermethylation, whereas other genes such as SLC6A4, DUSP6, and DTNBP1 are more likely to exhibit hypomethylation in promoter regions. In examining histone modifications, the literature suggests that clozapine changes histone methylation patterns in the prefrontal cortex, particularly elevating H3K4me3 at the Gad1 gene and affecting the transcription of genes like mGlu2 by modifying histone acetylation and interacting with HDAC2 enzymes. Risperidone and quetiapine, however, exhibit distinct impacts on histone marks across different brain regions and cell types, with risperidone reducing H3K27ac in the striatum and quetiapine modifying global H3K9me2 levels in the prefrontal cortex, suggesting antipsychotics demonstrate selective influence on histone modifications, which demonstrates a complex and targeted mode of action. While this review summarizes current knowledge, the intricate dynamics between antipsychotics and epigenetics clearly warrant more exhaustive exploration with the potential to redefine our understanding and treatment of psychiatric conditions. By deciphering the epigenetic changes associated with drug treatment and therapeutic outcomes, we can move closer to personalized medicine in psychiatry.