Role Of Histone Methyltransferases Research Articles

ARID5B promoted the histone demethylation of SORBS2 and hampered the metastasis of ovarian cancer

Ovarian cancer (OVCA) is the 4th most common female tumor after breast cancer, cervical cancer, and endometrial cancer, and now is mainly treated with debulking surgery and postoperative cisplatin and paclitaxel-based combination chemotherapy regimens. However, OVCA is insidious in its development and recurrence occurred in some patients after treatment. It is of great significance to study the pathogenesis of ovarian cancer and identify more biomarkers. Recently, the role of histone methyltransferase (HMT) and histone demethylase (HDM) in oncogenesis and development of malignant tumors has raised attention. Unlike other JMJC demethylases that have both JMJC and ARID domains in a single molecule, PHF2 requires assembly into a complex with a DNA-binding subunit (ARID5B) and exerts its enzymatic activity. Therefore, the aim of this manuscript is to investigate the role of histone demethylases ARID5B-PHF2 complex in the metastasis of OVCA. As result, we found ARID5B and PHF2 are both low expressed in OVCA tumor tissues and cell lines and associated with diagnosis and prognosis. Also, ARID5B suppressed rearrangement of the cytoskeleton in the process of EMT in OVCA cell lines. The role of PHF2 as a tumor suppressor was also confirmed both in vivo and in vitro. SORBS2 is low expressed in OVCA tumor tissues and cell lines and associated with diagnosis and prognosis. The expression of SORBS2 is positively corelated with the expression of ARID5B and PHF2. The promoter of SORBS2 is proved combined with ARID5B. The expression of SORBS2 was increased due to ARID5B-PHF2 complex promoted the histone demethylation by mainly binding in site H3K36me2 and therefore promoting the transcription of SORBS2. In conclusion, ARID5B-PHF2 complex promoted the histone demethylation of SORBS2 by mainly bind in site H3K36me2 and therefore promote the transcription of SORBS2 then hampered the process of EMT and tumor generation of OVCA. These results provided a new perspective on the molecular mechanisms of OVCA development and offered a new target of clinical diagnose and treatment of OVCA.

The role of histone methyltransferases in neurocognitive disorders associated with brain size abnormalities.

Brain size is controlled by several factors during neuronal development, including neural progenitor proliferation, neuronal arborization, gliogenesis, cell death, and synaptogenesis. Multiple neurodevelopmental disorders have co-morbid brain size abnormalities, such as microcephaly and macrocephaly. Mutations in histone methyltransferases that modify histone H3 on Lysine 36 and Lysine 4 (H3K36 and H3K4) have been identified in neurodevelopmental disorders involving both microcephaly and macrocephaly. H3K36 and H3K4 methylation are both associated with transcriptional activation and are proposed to sterically hinder the repressive activity of the Polycomb Repressor Complex 2 (PRC2). During neuronal development, tri-methylation of H3K27 (H3K27me3) by PRC2 leads to genome wide transcriptional repression of genes that regulate cell fate transitions and neuronal arborization. Here we provide a review of neurodevelopmental processes and disorders associated with H3K36 and H3K4 histone methyltransferases, with emphasis on processes that contribute to brain size abnormalities. Additionally, we discuss how the counteracting activities of H3K36 and H3K4 modifying enzymes vs. PRC2 could contribute to brain size abnormalities which is an underexplored mechanism in relation to brain size control.

Histone Methylation Is Required for Virulence, Conidiation, and Multi-Stress Resistance of Alternaria alternata

Histone methylation, which is critical for transcriptional regulation and various biological processes in eukaryotes, is a reversible dynamic process regulated by histone methyltransferases (HMTs) and histone demethylases (HDMs). This study determined the function of 5 HMTs (AaDot1, AaHMT1, AaHnrnp, AaSet1, and AaSet2) and 1 HDMs (AaGhd2) in the phytopathogenic fungus Alternaria alternata by analyzing targeted gene deletion mutants. The vegetative growth, conidiation, and pathogenicity of ∆AaSet1 and ∆AaSet2 were severely inhibited indicating that AaSet1 and AaSet2 play critical roles in cell development in A. alternata. Multiple stresses analysis revealed that both AaSet1 and AaSet2 were involved in the adaptation to cell wall interference agents and osmotic stress. Meanwhile, ∆AaSet1 and ∆AaSet2 displayed serious vegetative growth defects in sole carbon source medium, indicating that AaSet1 and AaSet2 play an important role in carbon source utilization. In addition, ∆AaSet2 colony displayed white in color, while the wild-type colony was dark brown, indicating AaSet2 is an essential gene for melanin biosynthesis in A. alternata. AaSet2 was required for the resistance to oxidative stress. On the other hand, all of ∆AaDot1, ∆AaHMT1, and ∆AaGhd2 mutants displayed wild-type phenotype in vegetative growth, multi-stress resistance, pathogenicity, carbon source utilization, and melanin biosynthesis. To explore the regulatory mechanism of AaSet1 and AaSet2, RNA-seq of these mutants and wild-type strain was performed. Phenotypes mentioned above correlated well with the differentially expressed genes in ∆AaSet1 and ∆AaSet2 according to the KEGG and GO enrichment results. Overall, our study provides genetic evidence that defines the central role of HMTs and HDMs in the pathological and biological functions of A. alternata.

Collateral Victim or Rescue Worker?-The Role of Histone Methyltransferases in DNA Damage Repair and Their Targeting for Therapeutic Opportunities in Cancer.

Disrupted DNA damage signaling greatly threatens cell integrity and plays significant roles in cancer. With recent advances in understanding the human genome and gene regulation in the context of DNA damage, chromatin biology, specifically biology of histone post-translational modifications (PTMs), has emerged as a popular field of study with great promise for cancer therapeutics. Here, we discuss how key histone methylation pathways contribute to DNA damage repair and impact tumorigenesis within this context, as well as the potential for their targeting as part of therapeutic strategies in cancer.

21 The contrasting role of histone methyltransferases during nuclear reprogramming: SUV39H knockdown improves bovine somatic cell nuclear transfer, while the absence of EHMT2 hampers it

The persistence of somatic epigenetic memory is indicated as the main barrier for efficient nuclear reprogramming. The augmented levels of histone methylation on somatic nuclei have been shown as the major regulator of this aberrant remodelling. Although they occupy the same site, H3K9me2 and H3K9me3 are catalysed by different histone methyltransferases (HMTs), EHMT2 and SUV39H, respectively. However, the influence of these HMTs during nuclear reprogramming is unclear. Hence, the main goal of this project was to investigate the role of HMTs responsible for H3K9 methylation during nuclear reprogramming and its consequences on embryo development. For this, we employed a small interfering RNA (siRNA)-mediated knockdown approach targeting EHMT2, SUV39H1, and SUV39H2 in bovine fetal fibroblasts. The RT-qPCR analyses showed ~80% reduction in total RNA after siRNA treatments for the target genes when compared with the control vector. We then quantified the H3K9me2 and H3K9me3 levels by immunostaining. The analysis displayed that H3K9me2 levels were diminished ~50% compared with control, whereas the reduction of H3K9me3 levels was only ~25%. Cells transfected with siRNA targeting EHMT2, SUV39H1, and SUV39H2 (All-siRNA) or control vector (control) were used as a nuclear donor on somatic cell nuclear transfer (SCNT) in five biological replicates. The IVF embryos were used as a biological control for immunostaining analysis. Embryos at both 8- to 16-cell and blastocysts stage (n=10 from 5 replicates) were collected to evaluate the effect of HMT knockdown on H3K9me2 and H3K9me3 levels by immunostaining. We found a reduction of blastocyst rates in the treatment 28±6.3 (mean±s.e.m.) when compared with control 60±4.8 (P=0.004). The immunostaining analysis showed that the levels of H3K9me2 and H3K9me3 were higher in cloned (control) than IVF embryos (P<0.05). Moreover, the All-siRNA group displayed a reduction in H3K9me2 levels compared with the control group and IVF through developmental stages analysed (P<0.05). Differently, H3K9me3 levels were higher in the All-siRNA group at the 8- to 16-cell stage, but no difference was found between treated and control groups at the blastocyst stage. We, therefore, decided to test whether individual knockdown would display a different result. We then used cells transfected with siRNA targeting only EHMT2 (EHMT2-siRNA), targeting SUV39H1 + SUV39H2 (SUV-siRNA), or control vector (control) as a nuclear donor on SCNT in five biological replicates. Surprisingly, the SUV-siRNA group increased blastocyst production 38±4.4 when compared with the control group 29±4.4 (P=0.01), whereas the EHMT2-siRNA showed a reduction in blastocyst rates: 21±5.6 (P=0.04). Our results indicate that EHMT2 has a key role during SCNT, possibly by its crosstalk with other modifications. Even though the SUV39H knockdown induced a small reduction in H3K9me3 levels in the nuclear donor, it was enough to increase the blastocyst rates by 10%. These results will allow us to better understand the complex mechanisms involved in the persistent epigenetic memory during nuclear reprogramming.

KMT2D deficiency enhances the anti-cancer activity of L48H37 in pancreatic ductal adenocarcinoma.

BACKGROUNDNovel therapeutic strategies are urgently needed for patients with a delayed diagnosis of pancreatic ductal adenocarcinoma (PDAC) in order to improve their chances of survival. Recent studies have shown potent anti-neoplastic effects of curcumin and its analogues. In addition, the role of histone methyltransferases on cancer therapeutics has also been elucidated. However, the relationship between these two factors in the treatment of pancreatic cancer remains unknown. Our working hypothesis was that L48H37, a novel curcumin analog, has better efficacy in pancreatic cancer cell growth inhibition in the absence of histone-lysine N-methyltransferase 2D (KMT2D).AIMTo determine the anti-cancer effects of L48H37 in PDAC, and the role of KMT2D on its therapeutic efficacy.METHODSThe viability and proliferation of primary (PANC-1 and MIA PaCa-2) and metastatic (SW1990 and ASPC-1) PDAC cell lines treated with L48H37 was determined by CCK8 and colony formation assay. Apoptosis, mitochondrial membrane potential (MMP), reactive oxygen species (ROS) levels, and cell cycle profile were determined by staining the cells with Annexin-V/7-AAD, JC-1, DCFH-DA, and PI respectively, as well as flow cytometric acquisition. In vitro migration was assessed by the wound healing assay. The protein and mRNA levels of relevant factors were analyzed using Western blotting, immunofluorescence and real time-quantitative PCR. The in situ expression of KMT2D in both human PDAC and paired adjacent normal tissues was determined by immunohistochemistry. In vivo tumor xenografts were established by injecting nude mice with PDAC cells. Bioinformatics analyses were also conducted using gene expression databases and TCGA.RESULTSL48H37 inhibited the proliferation and induced apoptosis in SW1990 and ASPC-1 cells in a dose- and time-dependent manner, while also reducing MMP, increasing ROS levels, arresting cell cycle at the G2/M stages and activating the endoplasmic reticulum (ER) stress-associated protein kinase RNA-like endoplasmic reticulum kinase/eukaryotic initiation factor 2α/activating transcription factor 4 (ATF4)/CHOP signaling pathway. Knocking down ATF4 significantly upregulated KMT2D in PDAC cells, and also decreased L48H37-induced apoptosis. Furthermore, silencing KMT2D in L48H37-treated cells significantly augmented apoptosis and the ER stress pathway, indicating that KMT2D depletion is essential for the anti-neoplastic effects of L48H37. Administering L48H37 to mice bearing tumors derived from control or KMT2D-knockdown PDAC cells significantly decreased the tumor burden. We also identified several differentially expressed genes in PDAC cell lines expressing very low levels of KMT2D that were functionally categorized into the extrinsic apoptotic signaling pathway. The KMT2D high- and low-expressing PDAC patients from the TCGA database showed similar survival rates,but higher KMT2D expression was associated with poor tumor grade in clinical and pathological analyses.CONCLUSIONL48H37 exerts a potent anti-cancer effect in PDAC, which is augmented by KMT2D deficiency.

The Role of Histone Methyltransferases and Long Non-coding RNAs in the Regulation of T Cell Fate Decisions.

T cell lineage decisions are critical for the development of proper immune responses to pathogens as well as important for the resolution of inflammatory responses. This differentiation process relies on a combination of intrinsic and extrinsic factors converging upon epigenetic regulation of transcriptional networks relevant to specific T cell lineages. As these biochemical modifications represent therapeutic opportunities in cancer biology and autoimmunity, implications of writers and readers of epigenetic marks to immune cell differentiation and function are highly relevant. Given the ready adoption of histone methyltransferase inhibitors in the clinic, we focus this review on the role of three histone modifying complexes: PRC-1, PRC-2, and G9A in modulating T cell fate decisions. Furthermore, we explore the role of long non-coding RNAs in regulating these processes, and discuss recent advances and challenges of implementing epigenetic therapies into clinical practice.

Epigenetics of cancer: the role of histone methyltransferase, SETDB1, in cancer metastasis

Cancer has a significant impact on society and the economy than all other disease, according to a report from the American Cancer Society.

Inhibition of histone methyltransferases SUV39H1 and G9a leads to neuroprotection in an in vitro model of cerebral ischemia.

Cerebral ischemia induces a complex transcriptional response with global changes in gene expression. It is essentially regulated by transcription factors as well as epigenetic players. While it is well known that the inhibition of transcriptionally repressive histone deacetylases leads to neuroprotection, the role of histone methyltransferases in the postischemic transcriptional response remains elusive. We investigated the effects of inhibition of the repressive H3K9 histone methyltransferases SUV39H1 and G9a on neuronal survival, H3K9 promoter signatures and gene expression. Their inhibition either with the specific blocker chaetocin or by use of RNA interference promoted neuronal survival in oxygen glucose deprivation (OGD). Brain-derived neurotrophic factor (BDNF) was upregulated and BDNF promoter regions showed an increase in histone marks characteristic for active transcription. The BDNF blockade with K252a abrogated the protective effect of chaetocin treatment. In conclusion, inhibition of histone methyltransferases SUV39H1 and G9a confers neuroprotection in a model of hypoxic metabolic stress, which is at least in part mediated by BDNF.

Exon resequencing of H3K9 methyltransferase complex genes, EHMT1, EHTM2 and WIZ, in Japanese autism subjects.

BackgroundHistone H3 methylation at lysine 9 (H3K9) is a conserved epigenetic signal, mediating heterochromatin formation by trimethylation, and transcriptional silencing by dimethylation. Defective GLP (Ehmt1) and G9a (Ehmt2) histone lysine methyltransferases, involved in mono and dimethylation of H3K9, confer autistic phenotypes and behavioral abnormalities in animal models. Moreover, EHMT1 loss of function results in Kleefstra syndrome, characterized by severe intellectual disability, developmental delays and psychiatric disorders. We examined the possible role of histone methyltransferases in the etiology of autism spectrum disorders (ASD) and suggest that rare functional variants in these genes that regulate H3K9 methylation may be associated with ASD.MethodsSince G9a-GLP-Wiz forms a heteromeric methyltransferase complex, all the protein-coding regions and exon/intron boundaries of EHMT1, EHMT2 and WIZ were sequenced in Japanese ASD subjects. The detected variants were prioritized based on novelty and functionality. The expression levels of these genes were tested in blood cells and postmortem brain samples from ASD and control subjects. Expression of EHMT1 and EHMT2 isoforms were determined by digital PCR.ResultsWe identified six nonsynonymous variants: three in EHMT1, two in EHMT2 and one in WIZ. Two variants, the EHMT1 ankyrin repeat domain (Lys968Arg) and EHMT2 SET domain (Thr961Ile) variants were present exclusively in cases, but showed no statistically significant association with ASD. The EHMT2 transcript expression was significantly elevated in the peripheral blood cells of ASD when compared with control samples; but not for EHMT1 and WIZ. Gene expression levels of EHMT1, EHMT2 and WIZ in Brodmann area (BA) 9, BA21, BA40 and the dorsal raphe nucleus (DoRN) regions from postmortem brain samples showed no significant changes between ASD and control subjects. Nor did expression levels of EHMT1 and EHMT2 isoforms in the prefrontal cortex differ significantly between ASD and control groups.ConclusionsWe identified two novel rare missense variants in the EHMT1 and EHMT2 genes of ASD patients. We surmise that these variants alone may not be sufficient to exert a significant effect on ASD pathogenesis. The elevated expression of EHMT2 in the peripheral blood cells may support the notion of a restrictive chromatin state in ASD, similar to schizophrenia.Electronic supplementary materialThe online version of this article (doi:10.1186/2040-2392-5-49) contains supplementary material, which is available to authorized users.

Involvement of histone methyltransferase GLP in HIV-1 latency through catalysis of H3K9 dimethylation

Understanding the mechanism of HIV-1 latency is crucial to eradication of the viral reservoir in HIV-1-infected individuals. However, the role of histone methyltransferase (HMT) G9a-like protein (GLP) in HIV-1 latency is still unclear. In the present work, we established four clonal cell lines containing HIV-1 vector. We found that the integration sites of most clonal cell lines favored active gene regions. However, we also observed hypomethylation of CpG of HIV 5′LTR in all four clonal cell lines. Additionally, 5′-deoxy-5′-methylthioadenosine (MTA), a broad-spectrum histone methyltransferase inhibitor, was used to examine the role of histone methylation in HIV-1 latency. MTA was found to decrease the level of H3K9 dimethylation, causing reactivation of latent HIV-1 in C11 cells. GLP knockdown by small interfering RNA clearly induced HIV-1 LTR expression. Results suggest that GLP may play a significant role in the maintenance of HIV-1 latency by catalyzing dimethylation of H3K9.

The Role of Histone Methyltransferases in Disease

The Role of Histone Methyltransferases in Disease