Mode of SUV420H2 heterochromatin localization through multiple HP1 binding motifs in the heterochromatic targeting module.

Constitutive heterochromatin is transcriptionally repressed and densely packed chromatin, typically harboring histone H3 Lys9 trimethylation (H3K9me3) and heterochromatin protein 1 (HP1). SUV420H2, a histone H4 Lys20 methyltransferase, is recruited to heterochromatin by binding to HP1 through its Heterochromatic Targeting Module (HTM). Here, we have identified three HP1 binding motifs within the HTM. Both the full-length HTM and its N-terminal region (HTM-N), which contains the first and second motifs, stabilized HP1 on heterochromatin. The intervening region between the first and second HP1 binding motifs in HTM-N was also crucial for HP1 binding. In contrast, the C-terminal region of HTM (HTM-C), containing the third motif, destabilized HP1 on chromatin. An HTM V374D mutant, featuring a Val374 to Asp substitution in the second HP1 binding motif, localizes to heterochromatin without affecting HP1 stability. These data suggest that the second HP1 binding motif in the SUV420H2 HTM is critical for locking HP1 on H3K9me3-enriched heterochromatin. HTM V374D, tagged with a fluorescent protein, can serve as a live-cell probe to visualize HP1-bound heterochromatin.

H3K9 and H4K20 methyltransferases are directly involved in the heterochromatinization of the paternal chromosomes in male Planococcus citri embryos.

Using a new method for bulk preparation of early stage embryos, we have investigated the role played by putative Planococcus citri H3K9 and H4K20 histone methyl transferases (HMTases) in regulating heterochromatinization of the imprinted paternal chromosomal set in male embryos. We found that H3K9 and H420 HMTases are required for heterochromatinization of the paternal chromosomes. We present evidence that both HMTases maintain the paternal "imprint" during the cleavage divisions when both parental chromosome sets are euchromatic. A testable model that accommodates our findings is proposed.

H4K20me3 marks distal intergenic and repetitive regions in human mature spermatozoa.

Sperm histones represent an essential part of the paternally transmitted epigenome, but uncertainty exists about the role of those remaining in non-coding and repetitive DNA. We therefore analyzed the genome-wide distribution of the heterochromatic marker H4K20me3 in human sperm and somatic (K562) cells. To specify the function of sperm histones, we compared all H4K20me3-containing and -free loci in the sperm genome. Sperm and somatic cells possessed a very similar H4K20me3 distribution: H4K20me3 peaks occurred mostly in distal intergenic regions and repetitive gene clusters (in particular genes encoding odorant-binding factors and zinc-finger antiviral proteins). In both cell types, H4K20me3 peaks were enriched in LINEs, ERVs, satellite DNA and low complexity repeats. In contrast, H4K20me3-free nucleosomes occurred more frequently in genic regions (in particular promoters, exons, 5'-UTR and 3'-UTR) and were enriched in genes encoding developmental factors (in particular transcription activators and repressors). H4K20me3-free nucleosomes were also detected in substantial quantities in distal intergenic regions and were enriched in SINEs. Thus, evidence suggests that paternally transmitted histones may have a dual purpose: maintenance and regulation of heterochromatin and guidance towards transcription of euchromatin.

H4K20me3 methyltransferase SUV420H2 shapes the chromatin landscape of pluripotent embryonic stem cells.

Heterochromatin, a densely packed chromatin state that is transcriptionally silent, is a critical regulator of gene expression. However, it is unclear how the repressive histone modification H4K20me3 or the histone methyltransferase SUV420H2 regulates embryonic stem (ES) cell fate by patterning the epigenetic landscape. Here, we report that depletion of SUV420H2 leads to a near-complete loss of H4K20me3 genome wide, dysregulated gene expression and delayed ES cell differentiation. SUV420H2-bound regions are enriched with repetitive DNA elements, which are de-repressed in SUV420H2 knockout ES cells. Moreover, SUV420H2 regulation of H4K20me3-marked heterochromatin controls chromatin architecture, including fine-scale chromatin interactions in pluripotent ES cells. Our results indicate that SUV420H2 plays a crucial role in stabilizing the three-dimensional chromatin landscape of ES cells, as loss of SUV420H2 resulted in A/B compartment switching, perturbed chromatin insulation, and altered chromatin interactions of pericentric heterochromatin and surrounding regions, indicative of localized decondensation. In addition, depletion of SUV420H2 resulted in compromised interactions between H4K20me3 and gene-regulatory regions. Together, these findings describe a new role for SUV420H2 in regulating the chromatin landscape of ES cells.

Combined low levels of H4K16ac and H4K20me3 predicts poor prognosis in breast cancer.

BACKGROUND: Results of previous studies about the prognostic roles of histone H4 lysine 16 acetylation (H4K16ac) and histone H4 lysine 20 trimethylation (H4K20me3) in breast cancer were inconsistent. Cellular experiments revealed the interplays between H4K16ac and H4K20me3, but no population study explored the interaction between them on the prognosis. METHODS: H4K16ac and H4K20me3 levels in tumors were evaluated by immunohistochemistry for 958 breast cancer patients. Hazard ratios for overall survival (OS) and progression-free survival (PFS) were estimated using Cox regression models. Interaction was assessed on multiplicative scale. Concordance index (C-index) was calculated to verify the predictive performance. RESULTS: The prognostic roles of the low level of H4K16ac or H4K20me3 were significant only in patients with the low level of another marker and their interactions were significant. Moreover, compared with joint high levels of both them, only the combined low levels of both them was associated with a poor prognosis but not the low level of single one. The C-index of the clinicopathological model combined the joint expression of H4K16ac and H4K20me3 [0.739 for OS; 0.672 for PFS] was significantly larger than that of the single clinicopathological model [0.699 for OS, P < 0.001; 0.642 for PFS, P = 0.003] or the model combined with the single H4K16ac [0.712 for OS, P < 0.001; 0.646 for PFS, P < 0.001] or H4K20me3 [0.724 for OS, P = 0.031; 0.662 for PFS, P = 0.006]. CONCLUSIONS: There was an interaction between H4K16ac and H4K20me3 on the prognosis of breast cancer and the combination of them was a superior prognostic marker compared to the single one.

KMT5c modulates adipocyte thermogenesis by regulating Trp53 expression.

Brown and beige adipocytes harbor the thermogenic capacity to adapt to environmental thermal or nutritional changes. Histone methylation is an essential epigenetic modification involved in the modulation of nonshivering thermogenesis in adipocytes. Here, we describe a molecular network leading by KMT5c, a H4K20 methyltransferase, that regulates adipocyte thermogenesis and systemic energy expenditure. The expression of Kmt5c is dramatically induced by a ß3-adrenergic signaling cascade in both brown and beige fat cells. Depleting Kmt5c in adipocytes in vivo leads to a decreased expression of thermogenic genes in both brown and subcutaneous (s.c.) fat tissues. These mice are prone to high-fat-diet-induced obesity and develop glucose intolerance. Enhanced transformation related protein 53 (Trp53) expression in Kmt5c knockout (KO) mice, that is due to the decreased repressive mark H4K20me3 on its proximal promoter, is responsible for the metabolic phenotypes. Together, these findings reveal the physiological role for KMT5c-mediated H4K20 methylation in the maintenance and activation of the thermogenic program in adipocytes.

Gene body methylation safeguards ribosomal DNA transcription by preventing PHF6-mediated enrichment of repressive histone mark H4K20me3.

DNA methylation shows complex correlations with gene expression, and the role of promoter hypermethylation in repressing gene transcription has been well addressed. Emerging evidence indicates that gene body methylation promotes transcription; however, the underlying mechanisms remain to be further investigated. Here, using methylated DNA immunoprecipitation sequencing (MeDIP-seq), bisulfite genomic sequencing, and immunofluorescent labeling, we show that gene body methylation is indeed positively correlated with rRNA gene (rDNA) transcription. Mechanistically, gene body methylation is largely maintained by DNA methyltransferase 1 (DNMT1), deficiency or downregulation of which during myoblast differentiation or nutrient deprivation results in decreased gene body methylation levels, leading to increased gene body occupancy of plant homeodomain (PHD) finger protein 6 (PHF6). PHF6 binds to hypomethylated rDNA gene bodies where it recruits histone methyltransferase SUV4-20H2 to establish the repressive histone modification, H4K20me3, ultimately inhibiting rDNA transcription. These findings demonstrate that DNMT1-mediated gene body methylation safeguards rDNA transcription by preventing enrichment of repressive histone modifications, suggesting that gene body methylation serves to maintain gene expression in response to developmental and/or environmental stresses.

The H4K20 methyltransferase Kmt5 is involved in secondary metabolism and stress response in phytopathogenic Fusarium species.

Fusarium fujikuroi and Fusarium graminearum are agronomically important plant pathogens, both infecting important staple food plants and thus leading to huge economic losses worldwide. F.fujikuroi belongs to the Fusarium fujikuroi species complex (FFSC) and causes bakanae disease on rice, whereas F.graminearum, a member of the Fusarium graminearum species complex (FGSC), is the causal agent of Fusarium Head Blight (FHB) disease on wheat, barley and maize. In recent years, the importance of chromatin regulation became evident in the plant-pathogen interaction. Several processes, including posttranslational modifications of histones, have been described as regulators of virulence and the biosynthesis of secondary metabolites. In this study, we have functionally characterised methylation of lysine 20 histone 4 (H4K20me) in both Fusarium species. We identified the respective genes solely responsible for H4K20 mono-, di- and trimethylation in F.fujikuroi (FfKMT5) and F.graminearum (FgKMT5). We show that loss of Kmt5 affects colony growth in F.graminearum while this is not the case for F.fujikuroi. Similarly, FgKmt5 is required for full virulence in F.graminearum as Δfgkmt5 is hypovirulent on wheat, whereas the F.fujikuroi Δffkmt5 strain did not deviate from the wild type during rice infection. Lack of Kmt5 had distinct effects on the secondary metabolism in both plant pathogens with the most pronounced effects on fusarin biosynthesis in F.fujikuroi and zearalenone biosynthesis in F.graminearum. Next to this, loss of Kmt5 resulted in an increased tolerance towards oxidative and osmotic stress in both species.

Maternal regulation of chromosomal imprinting in animals.

Chromosomal imprinting requires an epigenetic system that "imprints" one of the two parental chromosomes such that it results in a heritable (cell-to-cell) change in behavior of the "imprinted" chromosome. Imprinting takes place when the parental genomes are separate, which occurs during gamete formation in the respective germ-lines and post-fertilization during the period when the parental pro-nuclei lie separately within the ooplasm of the zygote. In the mouse, chromosomal imprinting is regulated by germ-line specific DNA methylation. But the methylation machinery in the respective germ-lines does not discriminate between imprinted and non-imprinted regions. As a consequence, the mouse oocyte nucleus contains over a thousand oocyte-specific germ-line differentially methylated regions (gDMRs). Upon fertilization, the sperm provides a few hundred sperm-specific gDMRs of its own. Combined, there are around 1600 imprinted and non-imprinted gDMRs in the pro-nuclei of the newly fertilized zygote. It is a remarkable fact that beginning in the maternal ooplasm, there are mechanisms that manage to preserve DNA methylation at ~ 26 known imprinted gDMRs in the face of the ongoing genome-wide DNA de-methylation that characterizes pre-implantation development. Specificity is achieved through the binding of KRAB-zinc finger proteins to their cognate recognition sequences within the gDMRs of imprinted genes. This in turn nucleates the assembly of localized heterochromatin-like complexes that preserve methylation at imprinted gDMRs through recruitment of the maintenance methyl transferase Dnmt1. These studies have shown that a germ-line imprint may cause parent-of-origin-specific behavior only if "licensed" by mechanisms that operate post-fertilization. Study of the germ-line and post-fertilization contributions to the imprinting of chromosomes in classical insect systems (Coccidae and Sciaridae) show that the ooplasm is the likely site where imprinting takes place. By comparing molecular and genetic studies across these three species, we suggest that mechanisms which operate post-fertilization play a key role in chromosomal imprinting phenomena in animals and conserved components of heterochromatin are shared by these mechanisms.

Tudor domain of histone demethylase KDM4B is a reader of H4K20me3.

The lysine histone demethylase KDM4B is overexpressed in several types of cancers and plays dual roles in genome stability maintenance. Although KDM4B is able to recognize several histone methylations, the underlying molecular mechanism is still unknown. In this study, we purified the KDM4B chromatin-associated hybrid tudor domains (HTDs) and plant home domains (PHDs) and performed the pull-down assay to screen the tri-methyl modified histone peptides that could be efficiently recognized by KDM4B. Our results showed that both HTD alone and the combination of HTD and PHD were able to specifically bind to H3K4me3 and H4K20me3. Because H4K20me3 is essential for KDM4B's rapid recruitment to DNA damage site, we further aligned the multiple tudor peptide sequence and identified two conserved residues Y993 and W987 that are critical for KDM4B-H4K20me3 interaction. The surface plasmon resonance analysis revealed that HTD displayed a rapid H4K20me3 bind-dissociate pattern. These findings therefore provided mechanistic insights into the binding of tudor domain of KDM4B protein with H4K20me3.

Identification of H4K20me3- and H3K4me3-associated RNAs using CARIP-Seq expands the transcriptional and epigenetic networks of embryonic stem cells.

RNA has been shown to interact with various proteins to regulate chromatin dynamics and gene expression. However, it is unknown whether RNAs associate with epigenetic marks such as post-translational modifications of histones, including histone 4 lysine 20 trimethylation (H4K20me3) or trimethylated histone 3 lysine 4 (H3K4me3), to regulate chromatin and gene expression. Here, we used chromatin-associated RNA immunoprecipitation (CARIP) followed by next-generation sequencing (CARIP-Seq) to survey RNAs associated with H4K20me3- and H3K4me3-marked chromatin on a global scale in embryonic stem (ES) cells. We identified thousands of mRNAs and noncoding RNAs that associate with H4K20me3- and H3K4me3-marked chromatin. H4K20me3- and H3K4me3-interacting RNAs are involved in chromatin organization and modification and RNA processing, whereas H4K20me3-only RNAs are involved in cell motility and differentiation, and H3K4me3-only RNAs are involved in metabolic processes and RNA processing. Expression of H3K4me3-associated RNAs is enriched in ES cells, whereas expression of H4K20me3-associated RNAs is enriched in ES cells and differentiated cells. H4K20me3- and H3K4me3-interacting RNAs originate from genes that co-localize with features of active chromatin, including transcriptional machinery and active promoter regions, and the histone modification H3K36me3 in gene body regions. We also found that H4K20me3 and H3K4me3 are associated with distinct gene features including transcripts of greater length and exon number relative to unoccupied transcripts. H4K20me3- and H3K4me3-marked chromatin is also associated with processed RNAs (exon transcripts) relative to unspliced pre-mRNA and ncRNA transcripts. In summary, our results provide evidence that H4K20me3- and H3K4me3-associated RNAs represent a distinct subnetwork of the ES cell transcriptional repertoire.

H3K9me3, H3K36me3, and H4K20me3 Expression Correlates with Patient Outcome in Esophageal Squamous Cell Carcinoma as Epigenetic Markers.

BACKGROUND: Histone methylation, as an essential pattern of posttranslational modifications, contributes to multiple cancer-related biological processes. Dysregulation of histone methylation is now considered a biomarker for cancer prognosis. AIMS: This study investigated and evaluated the potential role of four histone lysine trimethylation markers as biomarkers for esophageal squamous cell carcinoma (ESCC) prognosis. METHODS: Tissue arrays were made from 135 paraffin-embedded ESCC samples and examined for histone markers by immunohistochemistry, and 10 pairs of cancer and noncancerous mucosa tissues from ESCC patients were investigated with Western blot. Chi-squared test, Kaplan-Meier analysis with log-rank test, and Cox proportional hazard trend analyses were performed to assess the prognostic values of the markers. RESULTS: Histone 3 lysine 4 trimethylation (H3K4me3), histone 3 lysine 9 trimethylation (H3K9me3), and histone 4 lysine 20 trimethylation (H4K20me3), but not histone 3 lysine 36 trimethylation (H3K36me3), showed stronger immunostaining signals in tumor tissues than in the corresponding adjacent non-neoplastic mucosa tissues. The expression patterns of H3K36me3, H3K9me3, and H4K20me3 correlated with tumor infiltrating depth, lymph node involvement, and pTNM stage. Low-scoring H3K9me3 and H4K20me3 predicted better prognosis, while H3K36me3 manifested the opposite trend. Poor prognosis occurred in ESCC patients with expression patterns of high levels of H3K9me3, high levels of H4K20me3, and low levels of H3K36me3 expression. CONCLUSIONS: H3K9me3, H4K20me3, and H3K36me3 showed a close relationship with clinical features and were considered independent risk factors for survival of ESCC patients. The combination of H3K9me3, H4K20me3, and H3K36me3 expression, rather than the expression of a single histone marker, is believed to further enhance evaluations of ESCC prognosis and management.

Heterochromatic histone modifications at transposons in Xenopus tropicalis embryos.

Transposable elements are parasitic genomic elements that can be deleterious for host gene function and genome integrity. Heterochromatic histone modifications are involved in the repression of transposons. However, it remains unknown how these histone modifications mark different types of transposons during embryonic development. Here we document the variety of heterochromatic epigenetic signatures at parasitic elements during development in Xenopus tropicalis, using genome-wide ChIP-sequencing data and ChIP-qPCR analysis. We show that specific subsets of transposons in various families and subfamilies are marked by different combinations of the heterochromatic histone modifications H4K20me3, H3K9me2/3 and H3K27me3. Many DNA transposons are marked at the blastula stage already, whereas at retrotransposons the histone modifications generally accumulate at the gastrula stage or later. Furthermore, transposons marked by H3K9me3 and H4K20me3 are more prominent in gene deserts. Using intra-subfamily divergence as a proxy for age, we show that relatively young DNA transposons are preferentially marked by early embryonic H4K20me3 and H3K27me3. In contrast, relatively young retrotransposons are marked by increasing H3K9me3 and H4K20me3 during development, and are also linked to piRNA-sized small non-coding RNAs. Our results implicate distinct repression mechanisms that operate in a transposon-selective and developmental stage-specific fashion.

Heterochromatin Protein 1γ Is a Novel Epigenetic Repressor of Human Embryonic ϵ-Globin Gene Expression.

Production of hemoglobin during development is tightly regulated. For example, expression from the human ß-globin gene locus, comprising ß-, δ-, ϵ-, and γ-globin genes, switches from ϵ-globin to γ-globin during embryonic development and then from γ-globin to ß-globin after birth. Expression of human ϵ-globin in mice has been shown to ameliorate anemia caused by ß-globin mutations, including those causing ß-thalassemia and sickle cell disease, raising the prospect that reactivation of ϵ-globin expression could be used in managing these conditions in humans. Although the human globin genes are known to be regulated by a variety of multiprotein complexes containing enzymes that catalyze epigenetic modifications, the exact mechanisms controlling ϵ-globin gene silencing remain elusive. Here we found that the heterochromatin protein HP1γ, a multifunctional chromatin- and DNA-binding protein with roles in transcriptional activation and elongation, represses ϵ-globin expression by interacting with a histone-modifying enzyme, lysine methyltransferase SUV4-20h2. Silencing of HP1γ expression markedly decreased repressive histone marks and the multimethylation of histone H3 lysine 9 and H4 lysine 20, leading to a significant decrease in DNA methylation at the proximal promoter of the ϵ-globin gene and greatly increased ϵ-globin expression. In addition, using chromatin immunoprecipitation, we showed that SUV4-20h2 facilitates the deposition of HP1γ on the ϵ-globin-proximal promoter. Thus, these data indicate that HP1γ is a novel epigenetic repressor of ϵ-globin gene expression and provide a potential strategy for targeted therapies for ß-thalassemia and sickle cell disease.

H4K20me3 co-localizes with activating histone modifications at transcriptionally dynamic regions in embryonic stem cells.

BACKGROUND: Bivalent chromatin domains consisting of the activating histone 3 lysine 4 trimethylation (H3K4me3) and repressive histone 3 lysine 27 trimethylation (H3K27me3) histone modifications are enriched at developmental genes that are repressed in embryonic stem cells but active during differentiation. However, it is unknown whether another repressive histone modification, histone 4 lysine 20 trimethylation (H4K20me3), co-localizes with activating histone marks in ES cells. RESULTS: Here, we describe the previously uncharacterized coupling of the repressive H4K20me3 heterochromatin mark with the activating histone modifications H3K4me3 and histone 3 lysine 36 trimethylation (H3K36me3), and transcriptional machinery (RNA polymerase II; RNAPII), in ES cells. These newly described bivalent domains consisting of H3K4me3/H4K20me3 are predominantly located in intergenic regions and near transcriptional start sites of active genes, while H3K36me3/H4K20me3 are located in intergenic regions and within gene body regions of active genes. Global sequential ChIP, also termed reChIP-Seq, confirmed the simultaneous presence of H3K4me3 and H4K20me3 at the same genomic regions in ES cells. Genes containing H3K4me3/H4K20me3 exhibit decreased RNAPII pausing and are poised for deactivation of RNAPII binding during differentiation relative to H3K4me3 marked genes. An evaluation of transcription factor (TF) binding motif enrichment revealed that DNA sequence may play a role in shaping the landscape of these novel bivalent domains. Moreover, H3K4me3/H4K20me3 and H3K36me3/H4K20me3 bound regions are enriched with repetitive LINE and LTR elements. CONCLUSIONS: Overall, these findings highlight a previously undescribed subnetwork of ES cell transcriptional circuitry that utilizes dual marking of the repressive H4K20me3 mark with activating histone modifications.

FGF2 is a target and a trigger of epigenetic mechanisms associated with differences in emotionality: partnership with H3K9me3.

Posttranslational modifications of histone tails in chromatin template can result from environmental experiences such as stress and substance abuse. However, the role of epigenetic modifications as potential predisposing factors in affective behavior is less well established. To address this question, we used our selectively bred lines of high responder (bHR) and low responder (bLR) rats that show profound and stable differences in affective responses, with bLRs being prone to anxiety- and depression-like behavior and bHRs prone to addictive behavior. We first asked whether these phenotypes are associated with basal differences in epigenetic profiles. Our results reveal broad between-group differences in basal levels of trimethylated histone protein H3 at lysine 9 (H3K9me3) in hippocampus (HC), amygdala, and nucleus accumbens. Moreover, levels of association of H3K9me3 at Glucocorticoid Receptor (GR) and Fibroblast growth Factor 2 (FGF2) promoters differ reciprocally between bHRs and bLRs in these regions, consistent with these genes' opposing levels of expression and roles in modulating anxiety behavior. Importantly, this basal epigenetic pattern is modifiable by FGF2, a factor that modulates anxiety behavior. Thus, early-life FGF2, which decreases anxiety, altered the levels of H3K9me3 and its binding at FGF2 and GR promoters of bLRs rendering them more similar to bHRs. Conversely, knockdown of HC FGF2 altered both anxiety behavior and levels of H3K9me3 in bHRs, rendering them more bLR-like. These findings implicate FGF2 as a modifier of epigenetic mechanisms associated with emotional responsiveness, and point to H3K9me3 as a key player in the regulation of affective vulnerability.

SUV420H2 suppresses breast cancer cell invasion through down regulation of the SH2 domain-containing focal adhesion protein tensin-3.

The genome-wide loss of histone H4 lysine 20 tri-methylation (H4K20me3) is observed in multiple types of cancer, including breast tumors. Since H4K20me3 is preferentially targeted to repetitive elements in the pericentromeric and telomeric heterochromatin and plays a role in chromatin integrity, the pathological effects of disrupted H4K20me3 in tumors have been attributed to genomic instability. However, in this report, we show that loss of H4K20me3 modulates gene expression profiles, leading to increased cell invasion. Reduced H4K20me3 levels in tumor cells are often accompanied by a decrease in the expression of the H4K20-specific methyltransferase, SUV420H2. Exogenous delivery of SUV420H2 into MDA-MB-231 human breast cancer cells induced selective and specific changes in the expression of cancer-related genes. One of the most downregulated genes in response to SUV420H2 expression was the Src substrate, tensin-3, a focal adhesion protein that contributes to cancer cell migration. Depletion of tensin-3 suppressed breast cancer cell invasiveness. Furthermore, silencing of tensin-3 was associated with enrichment of H4K20me3 immediately upstream of the tensin-3 transcription start site, suggesting that the loss of H4K20me3 in tumor cells induced the expression of cancer-promoting genes. These findings connect the loss of H4K20me3 with tumor progression, through the transcriptional activation of cancer-promoting genes.

Histone 4 lysine 20 tri-methylation: a key epigenetic regulator in chromatin structure and disease.

Chromatin is a vital and dynamic structure that is carefully regulated to maintain proper cell homeostasis. A great deal of this regulation is dependent on histone proteins which have the ability to be dynamically modified on their tails via various post-translational modifications (PTMs). While multiple histone PTMs are studied and often work in concert to facilitate gene expression, here we focus on the tri-methylation of histone H4 on lysine 20 (H4K20me3) and its function in chromatin structure, cell cycle, DNA repair, and development. The recent studies evaluated in this review have shed light on how H4K20me3 is established and regulated by various interacting partners and how H4K20me3 and the proteins that interact with this PTM are involved in various diseases. Through analyzing the current literature on H4K20me3 function and regulation, we aim to summarize this knowledge and highlights gaps that remain in the field.

H4K20me3 upregulated by reactive oxygen species is associated with tumor progression and poor prognosis in patients with hepatocellular carcinoma.

Epigenetic alteration by oxidative stress is vitally involved in carcinogenesis and cancer progression. Previously, we demonstrated that oxidative stress was increased in hepatocellular carcinoma (HCC) patients and associated with tumor aggressiveness. Herein, we immunohistochemically investigated whether histone methylation, specifically H4K20me3, was upregulated in human hepatic tissues obtained from HCC patients (n = 100). Also, we experimentally explored if the H4K20me3 was upregulated by reactive oxygen species (ROS) and contributed to tumor progression in HCC cell lines. We found that H4K20me3 level was increased in HCC tissues compared with the adjacent noncancerous liver tissues. H3K9me3 and H3K4me3 levels were also increased in HCC tissues. Cox regression analysis revealed that the elevated H4K20me3 level was associated with tumor recurrence and short survival in HCC patients. Experimentally, H2O2 provoked oxidative stress and induced H4K20me3 formation in HepG2 and Huh7 cells. Transcript expression of histone methyltransferase Suv420h2 (for H4K20me3), Suv39h1 (for H3K9me3), and Smyd3 (for H3K4me3) were upregulated in H2O2-treated HCC cells. H2O2 also induced epithelial-mesenchymal transition (EMT) in HCC cells, indicated by decreased E-cadherin but increased α-SMA and MMP-9 mRNA expression. Migration, invasion, and colony formation in HCC cells were markedly increased following the H2O2 exposure. Inhibition of H4K20me3 formation by A196 (a selective inhibitor of Suv420h2) attenuated EMT and reduced tumor migration in H2O2-treated HCC cells. In conclusion, we demonstrated for the first time that H4K20me3 level was increased in human HCC tissues, and it was independently associated with poor prognosis in HCC patients. ROS upregulated H4K20me3 formation, induced mRNA expression of EMT markers, and promoted tumor progression in human HCC cells. Inhibition of H4K20me3 formation reduced EMT and tumor aggressive phenotypes in ROS-treated HCC cells. Possibly, ROS-induced EMT and tumor progression in HCC cells was epigenetically mediated through an increased formation of repressive chromatin H4K20me3.

A complex interplay between H2A.Z and HP1 isoforms regulates pericentric heterochromatin.

Pericentric heterochromatin (PCH) plays an essential role in the maintenance of genome integrity and alterations in PCH have been linked to cancer and aging. HP1 α, ß, and γ, are hallmarks of constitutive heterochromatin that are thought to promote PCH structure through binding to heterochromatin-specific histone modifications and interaction with a wide range of factors. Among the less understood components of PCH is the histone H2A variant H2A.Z, whose role in the organization and maintenance of PCH is poorly defined. Here we show that there is a complex interplay between H2A.Z and HP1 isoforms in PCH. While the loss of HP1α results in the accumulation of H2A.Z.1 in PCH, which is associated with a significant decrease in its mobile fraction, H2A.Z.1 binds preferentially to HP1ß in these regions. Of note, H2A.Z.1 downregulation results in increased heterochromatinization and instability of PCH, reflected by accumulation of the major epigenetic hallmarks of heterochromatin in these regions and increased frequency of chromosome aberrations related to centromeric/pericentromeric defects. Our studies support a role for H2A.Z in genome stability and unveil a key role of H2A.Z in the regulation of heterochromatin-specific epigenetic modifications through a complex interplay with the HP1 isoforms.