EZH2 promotes gastric cancer cells proliferation by repressing p21 expression

EZH2 is a core component of the polycomb repressive complex 2 (PRC2), which catalyzes trimethylation of histone H3 lysine 27 (H3K27me3) and promotes carcinogenesis by epigenetically silencing many tumor suppressor genes. Increased EZH2 expression is a marker of advanced and metastatic in many cancers, including lung, prostate and breast cancer, and it has been considered as a potential novel therapeutic target. However, the clinical significance and molecular mechanisms of EZH2 controlling gastric cancer cell proliferation and invasion are not well documented. In this study, immunohistochemical analysis was conducted to investigate the EZH2 expression in gastric cancer. We found that EZH2 levels were increased in gastric cancer tissues compared with adjacent normal tissues. Moreover, patients with high levels of EZH2 expression had a relatively poor prognosis. Furthermore, knockdown of EZH2 expression by siRNA could impair cell proliferation and invasion both in vitro and vivo. Finally, we found that EZH2 influences gastric cancer cells proliferation partly through regulating p21 expression. Our findings present that EZH2 over-expression can be identified as a poor prognostic biomarker in gastric cancer.     

Pharmacologic disruption of Polycomb-repressive complex 2-mediated gene repression selectively induces apoptosis in cancer cells

Polycomb-repressive complex 2 (PRC2)-mediated histone methylation plays an important role in aberrant cancer gene silencing and is a potential target for cancer therapy. Here we show that S-adenosylhomocysteine hydrolase inhibitor 3-Deazaneplanocin A (DZNep) induces efficient apoptotic cell death in cancer cells but not in normal cells. We found that DZNep effectively depleted cellular levels of PRC2 components EZH2, SUZ12, and EED and inhibited associated histone H3 Lys 27 methylation (but not H3 Lys 9 methylation). By integrating RNA interference (RNAi), genome-wide expression analysis, and chromatin immunoprecipitation (ChIP) studies, we have identified a prominent set of genes selectively repressed by PRC2 in breast cancer that can be reactivated by DZNep. We further demonstrate that the preferential reactivation of a set of these genes by DZNep, including a novel apoptosis affector, FBXO32, contributes to DZNep-induced apoptosis in breast cancer cells. Our results demonstrate the unique feature of DZNep as a novel chromatin remodeling compound and suggest that pharmacologic reversal of PRC2-mediated gene repression by DZNep may constitute a novel approach for cancer therapy.     

Weaver Syndrome‐Associated EZH2 Protein Variants Show Impaired Histone Methyltransferase Function In Vitro

Weaver syndrome (WS) is a rare congenital disorder characterized by generalized overgrowth, macrocephaly, specific facial features, accelerated bone age, intellectual disability, and susceptibility to cancers. De novo mutations in the enhancer of zeste homolog 2 (EZH2) have been shown to cause WS. EZH2 is a histone methyltransferase that acts as the catalytic agent of the polycomb‐repressive complex 2 (PRC2) to maintain gene repression via methylation of lysine 27 on histone H3 (H3K27). Functional studies investigating histone methyltransferase activity of mutant EZH2 from various cancers have been reported, whereas WS‐associated mutations remain poorly characterized. To investigate the role of EZH2 in WS, we performed functional studies using artificially assembled PRC2 complexes containing mutagenized human EZH2 that reflected the codon changes predicted from patients with WS. We found that WS‐associated amino acid alterations reduce the histone methyltransferase function of EZH2 in this in vitro assay. Our results support the hypothesis that WS is caused by constitutional mutations in EZH2 that alter the histone methyltransferase function of PRC2. However, histone methyltransferase activities of different EZH2 variants do not appear to correlate directly with the phenotypic variability between WS patients and individuals with a common c.553G>C (p.Asp185His) polymorphism in EZH2.     

The Polycomb Group Protein SUZ12 regulates histone H3 lysine 9 methylation and HP1α distribution

Regulation of histone methylation is critical for proper gene expression and chromosome function. Suppressor of Zeste 12 (SUZ12) is a requisite member of the EED/EZH2 histone methyltransferase complexes, and is required for full activity of these complexes in vitro. In mammals and flies, SUZ12/Su(z)12 is necessary for trimethylation of histone H3 on lysine 27 (H3K27me3) on facultative heterochromatin. However, Su(z)12 is unique among Polycomb Group Proteins in that Su(z)12 mutant flies exhibit gross defects in position effect variegation, suggesting a role for Su(z)12 in constitutive heterochromatin formation. We investigated the role of Suz12 in constitutive heterochromatin and discovered that Suz12 is required for histone H3 lysine 9 tri-methylation (H3K9me3) in differentiated but not undifferentiated mouse embryonic stem cells. Knockdown of SUZ12 in human cells caused a reduction in H3K27me3 and H3K9me3, and altered the distribution of HP1α. In contrast, EZH2 knockdown caused loss of H3K27me3 but not H3K9me3, indicating that SUZ12 regulates H3-K9 methylation in an EZH2-independent fashion. This work uncovers a role for SUZ12 in H3-K9 methylation.     

Evaluation of Histone 3 Lysine 27 Trimethylation (H3K27me3) and Enhancer of Zest 2 (EZH2) in Pediatric Glial and Glioneuronal Tumors Shows Decreased H3K27me3 in H3F3A K27M Mutant Glioblastomas

H3F3A mutations are seen in ～30% of pediatric glioblastoma (GBMs) and involve either the lysine residue at position 27 (K27M) or glycine at position 34 (G34R/V). Sixteen genes encode histone H3, each variant differing in only a few amino acids. Therefore, how mutations in a single H3 gene contribute to carcinogenesis is unknown. H3F3A K27M mutations are predicted to alter methylation of H3K27. H3K27me3 is a repressive mark critical to stem cell maintenance and is mediated by EZH2, a member of the polycomb‐group (PcG) family. We evaluated H3K27me3 and EZH2 expression using immunohistochemistry in 76 pediatric brain tumors. H3K27me3 was lowered/absent in tumor cells but preserved in endothelial cells and infiltrating lymphocytes in six out of 20 GBMs. H3K27me3 showed strong immunoreactivity in all other tumor subtypes. Sequencing of GBMs showed H3F3A K27M mutations in all six cases with lowered/absent H3K27me3. EZH2 expression was high in GBMs, but absent/focal in other tumors. However, no significant differences in EZH2 expression were observed between H3F3A K27M mutant and wild type GBMs, suggesting that EZH2 mediated trimethylation of H3K27 is inhibited in GBM harboring K27M mutations. Our results indicate that H3F3A K27M mutant GBMs show decreased H3K27me3 that may be of both diagnostic and biological relevance.     

Mutations in genes encoding polycomb repressive complex 2 subunits cause Weaver syndrome

Weaver syndrome (WS) is a rare congenital overgrowth disorder caused by heterozygous mutations in EZH2 (enhancer of zeste homolog 2) or EED (embryonic ectoderm development). EZH2 and EED are core components of the polycomb repressive complex 2 (PRC2), which possesses histone methyltransferase activity and catalyzes trimethylation of histone H3 at lysine 27. Here, we analyzed eight probands with clinically suspected WS by whole‐exome sequencing and identified three mutations: a 25.4‐kb deletion partially involving EZH2 and CUL1 (individual 1), a missense mutation (c.707G>C, p.Arg236Thr) in EED (individual 2), and a missense mutation (c.1829A>T, p.Glu610Val) in SUZ12 (suppressor of zeste 12 homolog) (individual 3) inherited from her father (individual 4) with a mosaic mutation. SUZ12 is another component of PRC2 and germline mutations in SUZ12 have not been previously reported in humans. In vitro functional analyses demonstrated that the identified EED and SUZ12 missense mutations cause decreased trimethylation of lysine 27 of histone H3. These data indicate that loss‐of‐function mutations of PRC2 components are an important cause of WS.     

EZH2在胚胎干细胞和肿瘤方面的研究进展

PcG蛋白家族成员EZH2是PCR2复合物的催化活性亚单位,通过甲基化核小体组蛋白H3的27位赖氨酸残基,沉默下游基因。这些下游靶基因大部分具有抑制肿瘤发生和调控干细胞分化的作用,它们的沉默将导致肿瘤的发生和干细胞多向分化潜能障碍。EZH2介导的基因沉默机制和DNA甲基转移酶(DNMTs)以及组蛋白脱乙酰酶(HDACs)之间存在密切的联系。现就EZH2在胚胎干细胞、肿瘤研究和临床应用前景作一综述。     

Methyltransferase expression and tumor suppressor gene methylation in sporadic and familial colorectal cancer

Molecular mechanisms underlying coordinated hypermethylation of multiple CpG islands in cancer remain unclear and studies of methyltransferase enzymes have arrived at conflicting results. We focused on DNMT1 and DNMT3B, DNA methyltransferases responsible for (de novo) methylation, and EZH2, histone (H3K27) methyltransferase, and examined their roles in tumor suppressor gene (TSG) methylation patterns we have previously established in sporadic and familial cancers. Our investigation comprised 165 tumors, stratified by tissue of origin (117 colorectal and 48 endometrial carcinomas) and sporadic vs. familial disease (57 sporadic vs. 60 familial, mainly Lynch syndrome, colorectal carcinomas). By immunohistochemical evaluation, EZH2 protein expression was associated with a TSG methylator phenotype. DNMT1, DNMT3B, and EZH2 were expressed at significantly higher levels in tumor vs. normal tissues. DNMT1 and EZH2 expression were positively correlated and higher in microsatellite‐unstable vs. microsatellite‐stable tumors, whether sporadic or hereditary. Ki‐67 expression mirrored the same pattern. Promoter methylation of the methyltransferase genes themselves was addressed as a possible cause behind their altered expression. While DNMT1 or EZH2 did not show differential methylation between normal and tumor tissues, DNMT3B analysis corroborated the regulatory role of a distal promoter region. Our study shows that methyltransferase expression in cancer depends on the tissue of origin, microsatellite‐instability status, cellular proliferation, and—in the case of DNMT3B—promoter methylation of the respective gene. Translation of methyltransferase expression into DNA methylation appears complex as suggested by the fact that except for EZH2, no clear association between methyltransferase protein expression and TSG methylation was observed. © 2015 Wiley Periodicals, Inc.     

PRC2‐complex related dysfunction in overgrowth syndromes: A review of EZH2, EED,and SUZ12 and their syndromic phenotypes

The EZH2, EED, and SUZ12 genes encode proteins that comprise core components of the polycomb repressive complex 2 (PRC2), an epigenetic “writer” with H3K27 methyltransferase activity, catalyzing the addition of up to three methyl groups on histone 3 at lysine residue 27 (H3K27). Partial loss‐of‐function variants in genes encoding the EZH2 and EED subunits of the complex lead to overgrowth, macrocephaly, advanced bone age, variable intellectual disability, and distinctive facial features. EZH2‐associated overgrowth, caused by constitutional heterozygous mutations within Enhancer of Zeste homologue 2 (EZH2), has a phenotypic spectrum ranging from tall stature without obvious intellectual disability or dysmorphic features to classical Weaver syndrome (OMIM #277590). EED‐associated overgrowth (Cohen–Gibson syndrome; OMIM #617561) is caused by germline heterozygous mutations in Embryonic Ectoderm Development (EED), and manifests overgrowth and intellectual disability (OGID), along with other features similar to Weaver syndrome. Most recently, rare coding variants in SUZ12 have also been described that present with clinical characteristics similar to the previous two syndromes. Here we review the PRC2 complex and clinical syndromes of OGID associated with core components EZH2, EED, and SUZ12.     

EZH2, new diagnosis and prognosis marker in acute myeloid leukemia patients

PurposeAcute myeloid leukemia (AML) is a heterogeneous disease. The discovery of novel discriminative biomarkers remains of utmost value for improving outcome predictions. Enhancer of zeste homolog 2 (EZH2) is a histone methyltransferase of H3K27me3. It is frequently up-regulated in human cancers and associated with silencing of differentiation genes. We aimed herein to investigate the prevalence and prognosis impact of somatic EZH2 mutations and their potential associations with other prognostic markers FLT3, NPM1, DNMT3A and IDH2.Materials and methodsOur study population was composed of 211 Tunisian patients with de novo AML and 14 healthy donors. The 11 last exons coding the set domain of EZH2 were investigated by PCR and Sanger sequencing.ResultsEZH2 mutations were identified in 66/211 (31%) patients with a sex ratio of 1.06. The presence of EZH2 mutations was statistically significantly associated with failure consolidation therapy (p = 0.004). There were no differences in the incidence of EZH2 mutations and FLT3-ITD, NPM1, DNMT3A and IDH2 mutations. When EZH2 mutations were associated with those of FLT3 or IDH2, a short duration of progression free survival was observed (p < 0.05). Moreover, CD7 aberrant markers conferred a poor prognosis in EZH2 mutated patients (p < 0.05).ConclusionsGiven these data we conclude that EZH2 mutations are frequent in our patients, and can be used as a prognosis marker in combination with FLT3, IDH2 mutations and CD7 marker, to stratify AML patients and to guide therapeutic decisions.