Suppressor Of Variegation 3-9 Homolog 1 Research Articles

Inhibition of SUV39H1 reduces tumor angiogenesis via Notch1 in oral squamous cell carcinoma

Inhibition of SUV39H1 reduces tumor angiogenesis via Notch1 in oral squamous cell carcinoma

SUV39H1 is a prognosis and immune microenvironment-related biomarker in diffuse large B-cell lymphoma.

The tumor microenvironment plays a crucial role in the oncogenesis and treatment of diffuse large B-cell lymphoma (DLBCL). The H3K9me3-specific histone methyltransferase Suppressor of variegation 3-9 homolog 1 (SUV39H1) is a significant gene that promotes the progression of various malignancies. However, the specific expression of SUV39H1 in DLBCL remains unclear. By retrieving data from GEPIA, UCSC XENA and TCGA public databases, we observed the high expression of SUV39H1 in DLBCL. Combined with an immunohistochemical validation assay, we analyzed our hospital's clinical characteristics and prognosis of 67 DLBCL patients. The results showed that high SUV39H1 expression was closely associated with age over 50years (P = 0.014) and low albumin levels (P = 0.023) of patients. Furthermore, the experiments in vitro were deployed to evaluate the regulation of SUV39H1 on the DLBCL immune microenvironment. The results showed that high SUV39H1 expression was closely associated with age over 50 years (P=0.014) and low albumin levels (P=0.023) of patients. The prognostic analysis showed that the high SUV39H1 expression group had a lower disease-free survival (DFS) rate than the low SUV39H1 expression group (P < 0.05). We further discovered that SUV39H1 upregulated the expression of CD86+ and CD163+ tumor-associated macrophages by DLBCL patients' tissues and cell experiments in vitro (P < 0.05). And SUV39H1-associated T lymphocyte subsets and cytokines IL-6/CCL-2 were downregulated in DLBCL (P < 0.05). In summary, SUV39H1 might be not only a potential target for treating DLBCL but also a clinical indicator for doctors to evaluate the trend of disease development.

SUV39H1 regulates corneal epithelial wound healing via H3K9me3-mediated repression of p27

BackgroundCorneal epithelial wound healing (CEWH) is vital for maintaining the integrity and barrier function of the cornea. Although histone modifications mediating gene expression patterns is fundamental in some other tissues, it remains unclear whether these gene regulation patterns underlie CEWH. Suppressor of variegation 3-9 homolog 1 (SUV39H1) plays a vital role in mediating gene silencing via histone H3 trimethylation of lysine 9 (H3K9me3). This study aims to characterize the comprehensive signature of epigenetic modifiers and determine the role of SUV39H1 in CEWH.MethodsNanoString nCounter technology was used to detect the differentially expressed epigenetic modifiers during CEWH. Bioinformatic analyses were performed to reveal their involvement in this process. After knockdown of SUV39H1 with siRNA transfection, we determined the function of SUV39H1 on cell proliferation and migration in human corneal epithelial cells (HCECs) via MTS, EdU, and wound-healing assay, respectively. Flow cytometry analysis further confirmed the effect of SUV39H1 on the cell cycle of HCECs. Loss-of-function assays for SUV39H1 with siRNA injection or chaetocin assessed the role of SUV39H1 on CEWH in vivo. Quantitative reverse transcription polymerase chain reaction (RT-qPCR) and Western blotting characterized the expression of SUV39H1 and its target genes. Chromatin immunoprecipitation assay was used to evaluate the distributions of H3K9me3 marks at the promoters of SUV39H1 target genes.ResultsWe first identified 92 differentially expressed epigenetic modifiers and revealed their involvement during CEWH. SUV39H1 was confirmed to be upregulated in response to corneal injury. Its downregulation significantly inhibited HCEC proliferation and retarded in vivo CEWH. Furthermore, knockdown of SUV39H1 upregulated the p27 expression level and reduced H3K9me3 marks at p27 promoter in HCECs. In addition, p27 was remarkably downregulated with elevated H3K9me3 marks at its promoter during in vivo CEWH.ConclusionsSUV39H1 plays a critical role in regulating corneal epithelial cell proliferation via H3K9me3-mediated suppression of p27 during CEWH. Our findings suggest that epigenetic modifiers such as SUV39H1 can be potential therapeutic approaches to accelerate corneal repair.

Differences in SUV39H1 and androgen receptor distribution in adenomyomatous hyperplasia and prostatic adenocarcinoma

Androgen receptor (AR) contributes to the growth of both early- and late-stage prostate cancer. Overexpression of suppressor of variegation 3-9 homolog 1 (SUV39H1) increases migration of prostate cancer cells, while depletion of SUV39H1 suppresses migration of prostate cancer cells. In this study, the aim was to show the relationships of AR and SUV39H1 with adenomyomatous hyperplasia (AH) and prostate adenocarcinoma (PCa). 70 AH and 70 PCa preparations in Pathology Department from 2013 to 16 were retrospectively investigated. Samples with immunohistochemical staining for AR and SUV39H1 were evaluated with a light microscope. After pathologic investigation of samples, AR and SUV39H1 expressions were scored. The changes in the frequencies of the obtained scores in the AH and PCa groups were analyzed statistically. AR expression was observed to be greater in AH compared to PCa. This difference was found to be statistically significant (p = 0.003). SUV39H1 expression was identified to be greater in PCa compared to AH and this showed statistical significance (p = 0.031). PCa samples were identified to have nearly 1.5 times more SUV39H1 mild staining compared to AH samples and this increase was two times for SUV39H1 strong staining. In our study, AR expression was greater in AH compared to PCa samples. This situation is inverse to the known mechanism and cannot be clearly explained. It needs to be supported with large series and other prognostic parameters. This study observed increased SUV39H1 values in PCa compared to AH and from this aspect, it may be considered an important poor prognosis parameter.

Phase-separated condensate-aided enrichment of biomolecular interactions for high-throughput drug screening in test tubes

Modification-dependent and -independent biomolecular interactions, including protein-protein, protein-DNA/RNA, protein-sugar, and protein-lipid interactions, play crucial roles in all cellular processes. Dysregulation of these biomolecular interactions or malfunction of the associated enzymes results in various diseases; therefore, these interactions and enzymes are attractive targets for therapies. High-throughput screening can greatly facilitate the discovery of drugs for these targets. Here, we describe a biomolecular interaction detection method, called phase-separated condensate-aided enrichment of biomolecular interactions in test tubes (CEBIT). The readout of CEBIT is the selective recruitment of biomolecules into phase-separated condensates harboring their cognate binding partners. We tailored CEBIT to detect various biomolecular interactions and activities of biomolecule-modifying enzymes. Using CEBIT-based high-throughput screening assays, we identified known inhibitors of the p53/MDM2 (MDM2) interaction and of the histone methyltransferase, suppressor of variegation 3-9 homolog 1 (SUV39H1), from a compound library. CEBIT is simple and versatile, and is likely to become a powerful tool for drug discovery and basic biomedical research.

Signal Activation of Hepatitis B Virus-Related Hepatocarcinogenesis by Up-regulation of SUV39h1.

Hepatitis B virus (HBV) X (HBx) protein is associated with hepatocellular carcinogenesis via the induction of malignant transformation and mitochondrial dysfunction. However, the association between HBx and histone methyltransferase in carcinogenesis has not been fully clarified. In the current study, we analyzed the association between HBx and the histone methyltransferase suppressor of variegation 3-9 homolog 1 (SUV39h1) using HBV replication models. We constructed several HBx and SUV39h1 expression plasmids and analyzed the association between HBx and SUV39h1 with respect to HBV replication and hepatocarcinogenesis. SUV39h1 up-regulation was observed in HBV-infected humanized mouse livers and clinical HBV-related hepatocellular carcinoma tissues, indicating that SUV39h1 expression might be regulated by HBV infection. Through in vitro analysis, we determined that the coactivator domain of HBx interacts with the PSET (PostSET) and SET (Su(var)3-9, Enhancer-of-zeste, Trithorax) domains of SUV39h1. The expression levels of 4 genes, activating transcription factor 6, α-fetoprotein, growth arrest and DNA damage-inducible 45a, and dual-specificity phosphatase 1, known to induce carcinogenesis via HBx expression, were up-regulated by HBx and further up-regulated in the presence of both HBx and SUV39h1. Furthermore, histone methyltransferase activity, the main function of SUV39h1, was enhanced in the presence of HBx. We demonstrated that SUV39h1 and HBx enhance each other's activity, leading to HBx-mediated hepatocarcinogenesis. We propose that regulation of this interaction could help suppress development of hepatocellular carcinoma.

Increase in hippocampal histone H3K9me3 is negatively correlated with memory in old male mice.

With advancing age, memory declines through different mechanisms including dysregulation of expression of synaptic plasticity genes in hippocampus. Increasing evidences suggest that these synaptic plasticity genes are regulated through epigenetic modifications. Recently we have reported that the neuronal immediate early genes (IEGs) are regulated by DNA methylation and histone acetylation, and their expression is downregulated in the hippocampus of old male mice, which subsequently results in decline of memory. These modifications do not work in isolation but act synergistically and lead to distinct regulation of gene expression. Therefore, in the present study, we have explored whether these genes are also regulated by histone methylation and this has any correlation with memory decline during aging. This study for the first time reports involvement of H3K9me3 in the regulation of neuronal IEGs during aging. Using novel object recognition and Y-maze test, the recognition and spatial memory was checked in male mice of different ages and it was found to decline in old. We have examined the expression of H3K9me3 specific histone methyltransferases and noted that only SUV39H1 (suppressor of variegation 3-9 homolog 1) increased significantly in old. Also the global H3K9me3 level was high in the hippocampus of old male mice. Further, chromatin immunoprecipitation assay revealed rise in H3K9me3 level at the promoter of IEGs in old as compared to young male mice. The immunofluorescence analysis also showed varying pattern of H3K9me3 expression in different subregions of hippocampus with aging. These findings showed negative correlation of increase in hippocampal histone H3K9me3 with memory decline in old male mice. Diagram here represents that during aging, there is increase in expression of SUV39H1. Such increased enzyme upregulates global and gene specific methylation in hippocampus of old male mice. H3K9me3 level increases at the promoter of neuronal IEGs leading to heterochromatisation and hence decrease in their expression and ultimately decline in memory during aging.

Contribution of the Suppressor of Variegation 3-9 Homolog 1 in Dorsal Root Ganglia and Spinal Cord Dorsal Horn to Nerve Injury–induced Nociceptive Hypersensitivity

Peripheral nerve injury-induced gene alterations in the dorsal root ganglion (DRG) and spinal cord likely participate in neuropathic pain genesis. Histone methylation gates gene expression. Whether the suppressor of variegation 3-9 homolog 1 (SUV39H1), a histone methyltransferase, contributes to nerve injury-induced nociceptive hypersensitivity is unknown. Quantitative real-time reverse transcription polymerase chain reaction analysis, Western blot analysis, or immunohistochemistry were carried out to examine the expression of SUV39H1 mRNA and protein in rat DRG and dorsal horn and its colocalization with DRG μ-opioid receptor (MOR). The effects of a SUV39H1 inhibitor (chaetocin) or SUV39H1 siRNA on fifth lumbar spinal nerve ligation (SNL)-induced DRG MOR down-regulation and nociceptive hypersensitivity were examined. SUV39H1 was detected in neuronal nuclei of the DRG and dorsal horn. It was distributed predominantly in small DRG neurons, in which it coexpressed with MOR. The level of SUV39H1 protein in both injured DRG and ipsilateral fifth lumbar dorsal horn was time dependently increased after SNL. SNL also produced an increase in the amount of SUV39H1 mRNA in the injured DRG (n = 6/time point). Intrathecal chaetocin or SUV39H1 siRNA as well as DRG or intraspinal microinjection of SUV39H1 siRNA impaired SNL-induced allodynia and hyperalgesia (n = 5/group/treatment). DRG microinjection of SUV39H1 siRNA also restored SNL-induced DRG MOR down-regulation (n = 6/group). The findings of this study suggest that SUV39H1 contributes to nerve injury-induced allodynia and hyperalgesia through gating MOR expression in the injured DRG. SUV39H1 may be a potential target for the therapeutic treatment of nerve injury-induced nociceptive hypersensitivity.

Stem Cell Depletion by Global Disorganization of the H3K9me3 Epigenetic Marker in Aging.

Epigenomic change and stem cell exhaustion are two of the hallmarks of aging. Accumulation of molecular damage is thought to underlie aging, but the precise molecular composition of the damage remains controversial. That some aging phenotypes, especially those that result from impaired stem cell function, are reversible suggest that such "damage" is repairable. Evidence is accumulating that dysfunction in aging stem cells results from increasing, albeit, subtle disorganization of the epigenome over time. Zhang et al. (2015) report that decreasing levels of WRN, Werner's syndrome (WS) helicase, with increasing age results in loss of heterochromatin marks in mesenchymal stem cells (MSCs) and correlates with an increased rate of cellular senescence. Although WRN plays a role in DNA repair, WRN exerted its effects on aging via maintaining heterochromatin, evidenced by reduced levels of interacting chromatin regulators heterochromatin protein 1α (HP1α), suppressor of variegation 3-9 homolog 1 (SUV39H1), and lamina-associated polypeptide 2β (LAP2β) as well as modified histone H3K9me3. Reducing expression of chromatin modeling co-factors SUV39H1 or HP1α in wild-type MSCs recapitulates the phenotype of WRN deficiency, resulting in reduced H3K9me3 levels and increased senescence without induction of markers of DNA damage, suggesting that chromatin disorganization and not DNA damage is responsible for the pathology of WS during aging in animals. Ectopic expression of HP1α restored H3K9me3 levels and repressed senescence in WRN-deficient MSCs. That HP1α can also suppress senescence in Hutchinson-Gilford progeria syndrome (HGPS) and extend life span in flies when over-expressed suggests that HP1α and H3K9me3 play conserved roles in maintenance of cell state. H3K9me3 levels are dynamic and expected to be potentially responsive to manipulation by extrinsic factors. Recent reports that migration inhibitory factor (MIF) or periodic fasting rejuvenate old MSCs provide the opportunity to link intrinsic and extrinsic mechanisms of aging in novel and potentially medically important ways and may lead to anti-aging treatments that reorganize the epigenome to rejuvenate cells and tissues.

Plant homeodomain finger protein 2 promotes bone formation by demethylating and activating Runx2 for osteoblast differentiation.

Plant homeodomain finger protein 2 (PHF2), which contains a plant homeodomain and a Jumonji-C domain, is an epigenetic regulator that demethylates lysine 9 in histone 3 (H3K9me2). On the other hand, runt-related transcription factor 2 (Runx2) plays essential roles in bone development and regeneration. Given previous reports that the PHF2 mutation can cause dwarfism in mice and that PHF2 expression is correlated with that of Runx2 in differentiating thymocytes, we investigated whether PHF2 regulates Runx2-mediated bone formation. Overexpression of PHF2 facilitated bone development in newborn mice, and viral shRNA-mediated knockdown of PHF2 delayed calvarial bone regeneration in adult rats. In primary osteoblasts and C2C12 precursor cells, PHF2 enhances osteoblast differentiation by demethylating Runx2, while suppressor of variegation 3-9 homolog 1 (SUV39H1) inhibits bone formation by methylating it. The PHF2-Runx2 interaction is mediated by the Jumonji-C and Runt domains of the two proteins, respectively. The interaction between Runx2 and osteocalcin promoter is regulated by the methylation status of Runx2, i.e., the interaction is augmented when Runx2 is demethylated. Our results suggest that SUV39H1 and PHF2 reciprocally regulate osteoblast differentiation by modulating Runx2-driven transcription at the post-translational level. This study may provide a theoretical basis for the development of new therapeutic modalities for patients with impaired bone development or delayed fracture healing.

Short-Chain Fatty Acids from Periodontal Pathogens Suppress Histone Deacetylases, EZH2, and SUV39H1 To Promote Kaposi's Sarcoma-Associated Herpesvirus Replication

Periodontal pathogens such as Porphyromonas gingivalis and Fusobacterium nucleatum produce five different short-chain fatty acids (SCFAs) as metabolic by-products. We detect significantly higher levels of SCFAs in the saliva of patients with severe periodontal disease. The different SCFAs stimulate lytic gene expression of Kaposi's sarcoma-associated herpesvirus (KSHV) dose dependently and synergistically. SCFAs inhibit class-1/2 histone deacetylases (HDACs) and downregulate expression of silent information regulator-1 (SIRT1). SCFAs also downregulate expression of enhancer of zeste homolog2 (EZH2) and suppressor of variegation 3-9 homolog1 (SUV39H1), which are two histone N-lysine methyltransferases (HLMTs). By suppressing the different components of host epigenetic regulatory machinery, SCFAs increase histone acetylation and decrease repressive histone trimethylations to transactivate the viral chromatin. These new findings provide mechanistic support that SCFAs from periodontal pathogens stimulate KSHV replication and infection in the oral cavity and are potential risk factors for development of oral Kaposi's sarcoma (KS). About 20% of KS patients develop KS lesions first in the oral cavity, while other patients never develop oral KS. It is not known if the oral microenvironment plays a role in oral KS tumor development. In this work, we demonstrate that a group of metabolic by-products, namely, short-chain fatty acids, from bacteria that cause periodontal disease promote lytic replication of KSHV, the etiological agent associated with KS. These new findings provide mechanistic support that periodontal pathogens create a unique microenvironment in the oral cavity that contributes to KSHV replication and development of oral KS.

Both histone methylation and acetylation contribute to the aberrant upregulation of proteinase 3 (PR3) and myeloperoxidase (MPO) genes in patients with ANCA disease

Histone lysine methylation (HKM) is an epigenetic change that establishes cell-specific gene expression and determines cell fates. In this study, we investigated the expression patterns of histone H3 lysine 9 methyltransferases (H3K9MTases) G9a (euchromatic histone lysine N-methyltransferase 2, Ehmt2), GLP (euchromatic histone lysine N-methyltransferase 1, Ehmt1), SETDB1 (SET domain, bifurcated 1), PRDM2 (PR domain containing 2), SUV39H1 (suppressor of variegation 3–9 homolog 1), and SUV39H2, as well as the distribution of 3 types of HKM at histone H3 lysine 9: mono- (H3K9me1), di- (H3K9me2), or tri-methylation (H3K9me3), during mouse growth plate development. In the forelimb cartilage primordial at embryonic day 12.5 (E12.5), none of the H3K9MTases were detected and H3K9me1, H3K9me2, and H3K9me3 were scarcely detected. At E14.5, the H3K9MTases were expressed at low levels in proliferating chondrocytes and at high levels in prehypertrophic and hypertrophic chondrocytes. Among the H3K9 methylations, H3K9me1 and H3K9me3 were markedly noted in these chondrocytes. At E16.5, G9, GLP, SETDB1, PRDM2, SUV39H1, and SUV39H2, as well as H3K9me1, H3K9me2, and H3K9me3, were detected in prehypertrophic and hypertrophic chondrocytes in the growth plate. Western blotting and real-time quantitative polymerase chain reaction analysis revealed the distributions of G9 and GLP proteins and the expression of all the H3K9MTase mRNAs in prehypertrophic and hypertrophic chondrocytes. These data suggest that H3K9 methyltransferases are predominantly expressed in prehypertrophic and hypertrophic chondrocytes, and that they could be involved in the regulation of gene expression and progression of chondrocyte differentiation by affecting the methylation state of histone H3 lysine 9 in the mouse growth plate.

Predominant expression of H3K9 methyltransferases in prehypertrophic and hypertrophic chondrocytes during mouse growth plate cartilage development

Histone lysine methylation (HKM) is an epigenetic change that establishes cell-specific gene expression and determines cell fates. In this study, we investigated the expression patterns of histone H3 lysine 9 methyltransferases (H3K9MTases) G9a (euchromatic histone lysine N-methyltransferase 2, Ehmt2), GLP (euchromatic histone lysine N-methyltransferase 1, Ehmt1), SETDB1 (SET domain, bifurcated 1), PRDM2 (PR domain containing 2), SUV39H1 (suppressor of variegation 3–9 homolog 1), and SUV39H2, as well as the distribution of 3 types of HKM at histone H3 lysine 9: mono- (H3K9me1), di- (H3K9me2), or tri-methylation (H3K9me3), during mouse growth plate development. In the forelimb cartilage primordial at embryonic day 12.5 (E12.5), none of the H3K9MTases were detected and H3K9me1, H3K9me2, and H3K9me3 were scarcely detected. At E14.5, the H3K9MTases were expressed at low levels in proliferating chondrocytes and at high levels in prehypertrophic and hypertrophic chondrocytes. Among the H3K9 methylations, H3K9me1 and H3K9me3 were markedly noted in these chondrocytes. At E16.5, G9, GLP, SETDB1, PRDM2, SUV39H1, and SUV39H2, as well as H3K9me1, H3K9me2, and H3K9me3, were detected in prehypertrophic and hypertrophic chondrocytes in the growth plate. Western blotting and real-time quantitative polymerase chain reaction analysis revealed the distributions of G9 and GLP proteins and the expression of all the H3K9MTase mRNAs in prehypertrophic and hypertrophic chondrocytes. These data suggest that H3K9 methyltransferases are predominantly expressed in prehypertrophic and hypertrophic chondrocytes, and that they could be involved in the regulation of gene expression and progression of chondrocyte differentiation by affecting the methylation state of histone H3 lysine 9 in the mouse growth plate.

Histone lysine methyltransferase, suppressor of variegation 3-9 homolog 1, promotes hepatocellular carcinoma progression and is negatively regulated by microRNA-125b

Hepatocellular carcinoma (HCC) is a major liver malignancy. We previously demonstrated that deregulation of epigenetic regulators is a common event in human HCC. Suppressor of variegation 3-9 homolog 1 (SUV39H1), the prototype of histone methyltransferase, is the major enzyme responsible for histone H3 lysine 9 trimethylation, which, essentially, is involved in heterochromatin formation, chromosome segregation, and mitotic progression. However, the implication of SUV39H1 in hepatocarcinogenesis remains elusive. In this study, we found that SUV39H1 was frequently up-regulated in human HCCs and was significantly associated with increased Ki67 expression (P < 0.001) and the presence of venous invasion (P = 0.017). To investigate the role of SUV39H1 in HCC development, both gain- and loss-of-function models were established. SUV39H1 overexpression remarkably enhanced HCC cell clonogenicity, whereas knockdown of SUV39H1 substantially suppressed HCC cell proliferation and induced cell senescence. In addition, ectopic expression of SUV39H1 increased the migratory ability of HCC cells, whereas a reduced migration rate was observed in SUV39H1 knockdown cells. The significance of SUV39H1 in HCC was further demonstrated in a nude mice model; SUV39H1 knockdown drastically inhibited in vivo tumorigenicity and abolished pulmonary metastasis of HCC cells. We also identified microRNA-125b (miR-125b) as a post-transcriptional regulator of SUV39H1. Ectopic expression of miR-125b inhibited SUV39H1 3'-untranslated-region-coupled luciferase activity and suppressed endogenous SUV39H1 expression at both messenger RNA and protein levels. We have previously reported frequent down-regulation of miR-125b in HCC. Interestingly, miR-125b level was found to be inversely correlated with SUV39H1 expression (P = 0.001) in clinical specimens. Our observations suggested that miR-125b down-regulation may account for the aberrant SUV39H1 level in HCC. Our study demonstrated that SUV39H1 up-regulation contributed to HCC development and metastasis. The tumor-suppressive miR-125b served as a negative regulator of SUV39H1.

Epigenetic modifications and related mRNA expression during bovine oocyte in vitro maturation

The present study investigated the global pattern of two histone modifications and methylation of DNA during in vitro maturation of bovine oocytes retrieved from follicles of two different sizes (<2 mm and 2-8 mm). The methylation status of histone H3 at position lysine K9 (H3K9 me2), the acetylation status of histone H4 at position lysine K12 (H4K12ac) and the methylation of DNA were assessed by immunocytochemistry. In parallel, the relative abundance of mRNAs coding for proteins specifically involved in reprogramming, including HLA-B associated transcript 8 (G9A), suppressor of variegation 3-9 homolog 1 (SUV39H1), the somatic isoform of DNA methyltransferase 1 (DNMT1), DNA methyltransferase 3b (DNMT3b) and zygote arrest 1 (ZAR1) was determined by RT-PCR. The alpha-H3K9 me2 signal was present in the GV stage and remained detectable until the end of the maturation period. alpha-H4K12ac antibody gave a stronger signal in GV and GVBD oocytes and markedly decreased after GVBD. The signal showing the methylation of DNA was present during the entire maturation period. The five transcripts showed a gene-specific expression profile. Results revealed the global patterns of H3K9 me2, H4K12ac, DNA methylation and the mRNA pool profiles of genes critically involved in epigenetic modifications during bovine oocyte maturation and their possible relationship with the acquisition of oocyte developmental competence and follicular development.

EVI1 Is a Transcriptional Repressor Able To Recruit SUV39H1 Histone Methylation Activity.

The Evi1 (Ecotropic viral integration site 1) gene is located on chromosome 3q26 and encodes a highly conserved nuclear protein that belongs to the Kruppel family of transcription factors, containing two Cys2-His2 repeat zinc finger motifs domains. EVI1 is not expressed in normal hematopoietic tissues, but can be found overexpressed in acute myeloid leukemia/myelodysplastic syndrome (AML/MDS) cells as a consequence of chromosome 3 rearrangements. In vitro and in vivo studies have shown that a number of biological properties can be attributed to the EVI1 protein that might contribute to leukemogenesis: deregulation of cell proliferation, inhibition of TGF-ß signaling pathway, and suppression of stress induced apoptosis. We and others reported that EVI1 acts as a transcriptional repressor through its interaction with a subset of co-repressor proteins. More recently, we showed that EVI1 directly interacts with RUNX1 and GATA1 transcription factors. In both cases this interaction alters the ability of the transcription factors to bind their recognition sites on the DNA, leading to the deregulation of their target genes and to the impairment of the normal hematopoietic development. Interestingly, EVI1-mediated transcriptional silencing in association with HDAC proteins can be only partially rescued by HDAC inhibitors, suggesting that additional chromosomal modification might occur for gene silencing. Methylation by histone methyltransferase proteins (HMTs) is one type of these modifications, and when it occurs on certain residues can lead both to repression or activation of gene expression. The SUV39H1 (Suppressor of Variegation 3–9 Homolog 1) gene encodes a histone 3 specific methyltransferase. SUV39H1 selectively methylates lysine 9 of the amino terminus of the histone 3. Tri-methylation of histone H3 is important for recruiting heterochromatin protein 1 (HP1), thereby regulating gene expression, chromatin packaging and heterochromatin formation. Here we investigate the hypothesis that histone methylation may contribute to the transcriptional repressive potential of EVI1. We found that EVI1 physically interacts with the histone methyltransferase SUV39H1. The interaction between the two proteins requires the N-terminus of EVI1 where the proximal zinc finger domain is located; in particular, out of the seven motifs that compose this domain, the distal three motifs seem to be responsible for the binding between the two proteins. By pull-down assay with in vitro translated EVI1 this interaction appears to be direct. Futhermore, using in vitro histone methylation assay we show that EVI1 forms a complex with SUV39H1 that is able to methylate a recombinant H3, suggesting that the EVI1-SUV39H1 interaction does not affect the histone methyltransferase activity of SUV39H1. We suggest here that EVI1 interacts with SUV39H1 and that this binding could have a role in the epigenetic remodellig of the chromatin. According to this model, it is possible that EVI1 recruits histone methylation activity to selected genomic loci to silence tumor suppressor genes and repress differentiation factors. Because lysine methylation by SUV39H1 cannot take place on an already acetylated lysine, deacetylation activity associated with EVI1, through its interaction with HDACs, becomes a necessary preceeding step to SUV39H1 mediated histone methylation.