Histone H4 asymmetrically dimethylated at arginine 3 (H4R3me2a), a mark of super-enhancers.

Histone H4 asymmetrically dimethylated at arginine 3 (H4R3me2a) is an active histone mark catalyzed by protein arginine methyltransferase 1 (PRMT1), a major arginine methyltransferase in vertebrates catalyzing asymmetric dimethylation of arginine. H4R3me2a stimulates the activity of lysine acetyltransferases such as CBP/p300, which catalyze the acetylation of H3K27, a mark of active enhancers, super-enhancers, and promoters. There are a few studies on the genomic location of H4R3me2a. In chicken polychromatic erythrocytes, H4R3me2a is found in introns and intergenic regions and binds to the globin locus control region (a super-enhancer) and globin regulatory regions. In this report, we analyzed chromatin immunoprecipitation sequencing data for the genomic location of H4R3me2a in the breast cancer cell line MCF7. As in avian cells, MCF7 H4R3me2a is present in intronic and intergenic regions. Nucleosomes with H4R3me2a and H3K27ac next to nucleosome-free regions are found at super-enhancers, enhancers, and promoter regions of expressed genes. Genes with critical roles in breast cancer cells have broad domains of nucleosomes with H4R3me2a, H3K27ac, and H3K4me3. Our results are consistent with PRMT1-mediated H4R3me2a playing a key role in the chromatin organization of regulatory regions of vertebrate genomes.

Neurodevelopmental functions and activities of the KAT3 class of lysine acetyltransferases.

The human lysine acetyltransferases KAT3A (CREBBP) and KAT3B (EP300) are essential enzymes in gene regulation in the nucleus. Their ubiquitous expression in metazoan cell types controls cell proliferation and differentiation during development. This comprehensive review delves into the biological roles of KAT3A and KAT3B in neurodevelopment, shedding light on how alterations in their regulation or activity can potentially contribute to a spectrum of neurodegenerative diseases (e.g., Huntington's and Alzheimer's). We explore the pathophysiological implications of KAT3 function loss in these disorders, considering their conserved protein domains and biochemical functions in chromatin regulation. The discussion also underscores the crucial role of KAT3 proteins and their substrates in supporting the integration of key cell signaling pathways. Furthermore, the narrative highlights the interdependence of KAT3-mediated lysine acetylation with lysine methylation and arginine methylation. From a cellular perspective, KAT3-dependent signal integration at subnuclear domains is mediated by liquid-liquid phase separation in response to KAT3-mediated lysine acetylation. The disruption of these finely tuned regulatory processes underscores their pathological roles in neurodegeneration. This review also points to the exciting potential for future research in this field, inspiring further investigation and discovery in the area of neurodevelopment and neurodegenerative diseases.

Protein arginine methyltransferase 1, a major regulator of biological processes.

Protein arginine methyltransferase 1 (PRMT1) is a major type I arginine methyltransferase that catalyzes the formation of monomethyl and asymmetric dimethylarginine in protein substrates. It was first identified to asymmetrically methylate histone H4 at the third arginine residue forming the H4R3me2a active histone mark. However, several protein substrates are now identified as being methylated by PRMT1. As a result of its association with diverse classes of substrates, PRMT1 regulates several biological processes like chromatin dynamics, transcription, RNA processing, and signal transduction. The review provides an overview of PRMT1 structure, biochemical features, specificity, regulation, and role in cellular functions. We discuss the genomic distribution of PRMT1 and its association with tRNA genes. Further, we explore the different substrates of PRMT1 involved in splicing. In the end, we discuss the proteins that interact with PRMT1 and their downstream effects in diseased states.

Mitogen- and stress-activated protein kinase (MSK1/2) regulated gene expression in normal and disease states.

The mitogen- and stress-activated protein kinases (MSK) are epigenetic modifiers that regulate gene expression in normal and disease cell states. MSK1 and 2 are involved in a chain of signal transduction events bringing signals from the external environment of a cell to specific sites in the genome. MSK1/2 phosphorylate histone H3 at multiple sites, resulting in chromatin remodeling at regulatory elements of target genes and the induction of gene expression. Several transcription factors (RELA of NF-κB and CREB) are also phosphorylated by MSK1/2 and contribute to induction of gene expression. In response to signal transduction pathways, MSK1/2 can stimulate genes involved in cell proliferation, inflammation, innate immunity, neuronal function, and neoplastic transformation. Abrogation of the MSK-involved signaling pathway is among the mechanisms by which pathogenic bacteria subdue the host's innate immunity. Depending on the signal transduction pathways in play and the MSK-targeted genes, MSK may promote or hinder metastasis. Thus, depending on the type of cancer and genes involved, MSK overexpression may be a good or poor prognostic factor. In this review, we focus on mechanisms by which MSK1/2 regulate gene expression, and recent studies on their roles in normal and diseased cells.

Broad histone H4 monomethylation marks expressed genes involved in translation.

H4K20me1 (histone H4 monomethylated at lysine 20) generally has a broad distribution along genes and has been reported to be associated with expressed and repressed genes. In contrast, H3K4me3 (histone H3 trimethylated at lysine 4) is positioned as a narrow peak at the 5' end of most expressed genes in vertebrate cells. A small population of genes involved in cell identity has H3K4me3 distributed throughout the gene body. In this report, we show that H4K20me1 is associated with expressed genes in estrogen receptor-positive breast cancer MCF7 cells and erythroleukemic K562 cells. Further, we identified the genes with the broadest H4K20me1 domains in these two cell types. The broad H4K20me1 domain marked gene bodies of expressed genes, but not the promoter or enhancer regions. The most significant GO term (biological processes) of these genes was cytoplasmic translation. There was little overlap between the genes marked with the broad H4K20me1 domain and those marked with H3K4me3. H4K20me1 and H3K79me2 distributions along expressed gene bodies were similar, suggesting a relationship between the enzymes catalyzing these histone modifications.

Chicken Erythrocyte: Epigenomic Regulation of Gene Activity.

The chicken genome is one-third the size of the human genome and has a similarity of sixty percent when it comes to gene content. Harboring similar genome sequences, chickens' gene arrangement is closer to the human genomic organization than it is to rodents. Chickens have been used as model organisms to study evolution, epigenome, and diseases. The chicken nucleated erythrocyte's physiological function is to carry oxygen to the tissues and remove carbon dioxide. The erythrocyte also supports the innate immune response in protecting the chicken from pathogens. Among the highly studied aspects in the field of epigenetics are modifications of DNA, histones, and their variants. In understanding the organization of transcriptionally active chromatin, studies on the chicken nucleated erythrocyte have been important. Through the application of a variety of epigenomic approaches, we and others have determined the chromatin structure of expressed/poised genes involved in the physiological functions of the erythrocyte. As the chicken erythrocyte has a nucleus and is readily isolated from the animal, the chicken erythrocyte epigenome has been studied as a biomarker of an animal's long-term exposure to stress. In this review, epigenomic features that allow erythroid gene expression in a highly repressive chromatin background are presented.

Differential expression of HNF1A and HNF1A-AS1 in colon cancer cells.

The human hepatocyte nuclear factor 1 homeobox A (HNF1A) gene loci express the protein-coding HNF1A transcript and a long non-coding RNA in the anti-sense (HNF1A-AS1) direction. HNF1A-AS1 is expressed in numerous types of cancers and poor clinical outcomes such as higher mortality rates, greater metastatic capacity, and poor prognosis of the disease are the results of this expression. In this study, we determined the epigenetic features of the HNF1A gene loci, and expression and cellular localization of HNF1A-AS1 RNA, HNF1A RNA, and HNF1A protein in colorectal cancer (HT-29, HTC116, RKO, and SW480) and normal colon epithelial (CCD841) cells. The HT-29 HNF1A gene had active histone marks (H3K4me3, H3K27ac) and DNase 1 accessible sites at the promoter regions of the HNF1A and HNF1A-AS1 genes. These epigenetic marks were not observed in the other colorectal cancer cells or in the normal colon epithelial cells. Consistent with the active gene epigenetic signature of the HNF1A gene in HT-29 cells, HNF1A protein, and HNF1A/HNF1A-AS1 transcripts were detected in HT-29 cells but poorly, if at all observed, in the other cell types. In HT-29 cells, HNF1A-AS1 localized to the nucleus and was found to bind to the enhancer of zeste homolog 2 (EZH2, a member of PRC2 complex) and potentially form RNA-DNA triplexes with DNase 1 accessible sites in the HT-29 genome. These activities of HNF1A-AS1 may contribute to the oncogenic properties of this long non-coding RNA.

The Chromatin Structure at the MECP2 Gene and In Silico Prediction of Potential Coding and Non-Coding MECP2 Splice Variants.

Methyl CpG binding protein 2 (MeCP2) is an epigenetic reader that binds to methylated CpG dinucleotides and regulates gene transcription. Mecp2/MECP2 gene has 4 exons, encoding for protein isoforms MeCP2E1 and MeCP2E2. MeCP2 plays key roles in neurodevelopment, therefore, its gain- and loss-of-function mutations lead to neurodevelopmental disorders including Rett Syndrome. Here, we describe the structure, functional domains, and evidence support for potential additional alternatively spliced MECP2 transcripts and protein isoforms. We conclude that NCBI MeCP2 isoforms 3 and 4 contain certain MeCP2 functional domains. Our in silico analysis led to identification of histone modification and accessibility profiles at the MECP2 gene and its cis-regulatory elements. We conclude that the human MECP2 gene associated histone post-translational modifications exhibit high similarity between males and females. Between brain regions, histone modifications were found to be less conserved and enriched within larger genomic segments named as "S1-S11". We also identified highly conserved DNA accessibility regions in different tissues and brain regions, named as "A1-A9" and "B1-B9". DNA methylation profile was similar between mid-frontal gyrus of donors 35 days-25 years of age. Based on ATAC-seq data, the identified hypomethylated regions "H1-H8" intersected with most regions of the accessible chromatin (A regions).

The chicken model organism for epigenomic research.

The chicken model organism has advanced the areas of developmental biology, virology, immunology, oncology, epigenetic regulation of gene expression, conservation biology, and genomics of domestication. Further, the chicken model organism has aided in our understanding of human disease. Through the recent advances in high-throughput sequencing and bioinformatic tools, researchers have successfully identified sequences in the chicken genome that have human orthologs, improving mammalian genome annotation. In this review, we highlight the importance of chicken as an animal model in basic and pre-clinical research. We will present the importance of chicken in poultry epigenetics and in genomic studies that trace back to their ancestor, the last link between human and chicken in the tree of life. There are still many genes of unknown function in the chicken genome yet to be characterized. By taking advantage of recent sequencing technologies, it is possible to gain further insight into the chicken epigenome.

Epigenetic regulation of ACE2, the receptor of the SARS-CoV-2 virus1.

The angiotensin-converting enzyme 2 (ACE2) is the receptor for the three coronaviruses HCoV-NL63, SARS-CoV, and SARS-CoV-2. ACE2 is involved in the regulation of the renin-angiotensin system and blood pressure. ACE2 is also involved in the regulation of several signaling pathways, including integrin signaling. ACE2 expression is regulated transcriptionally and post-transcriptionally. The expression of the gene is regulated by two promoters, with usage varying among tissues. ACE2 expression is greatest in the small intestine, kidney, and heart and detectable in a variety of tissues and cell types. Herein we review the chemical and mechanical signal transduction pathways regulating the expression of the ACE2 gene and the epigenetic/chromatin features of the expressed gene.

The treatment of SARS-CoV2 with antivirals and mitigation of the cytokine storm syndrome: the role of gene expression.

In the absence of a vaccine, the treatment of SARS-CoV2 has focused on eliminating the virus with antivirals or mitigating the cytokine storm syndrome (CSS) that leads to the most common cause of death: respiratory failure. Herein we discuss the mechanisms of antiviral treatments for SARS-CoV2 and treatment strategies for the CSS. Antivirals that have shown in vitro activity against SARS-CoV2, or the closely related SARS-CoV1 and MERS-CoV, are compared on the enzymatic level and by potency in cells. For treatment of the CSS, we discuss medications that reduce the effects or expression of cytokines involved in the CSS with an emphasis on those that reduce IL-6 because of its central role in the development of the CSS. We show that some of the medications covered influence the activity or expression of enzymes involved in epigenetic processes and specifically those that add or remove modifications to histones or DNA. Where available, the latest clinical data showing the efficacy of the medications is presented. With respect to their mechanisms, we explain why some medications are successful, why others have failed, and why some untested medications may yet prove useful.

Atypical chromatin structure of immune-related genes expressed in chicken erythrocytes.

The major biological role of red blood cells is to carry oxygen to the tissues in the body. However, another role of the erythroid cell is to participate in the immune response. Mature erythrocytes from chickens express Toll-like receptors and several cytokines in response to stimulation of the immune system. We previously reported the application of a biochemical fractionation protocol to isolate highly enriched transcribed DNA from polychromatic erythrocytes from chickens. In conjunction with next-generation DNA, RNA sequencing, chromatin immunoprecipitation-DNA sequencing, and formaldehyde-assisted isolation of regulatory elements (FAIRE) sequencing, we identified the active chromosomal compartments and determined their structural signatures in relation to expression levels. Here, we present the detailed chromatin characteristics of erythroid genes participating in the innate immune response. Our studies revealed an atypical chromatin structure for several genes coding for Toll-like receptors, interleukins, and interferon regulatory factors. The body of these genes had nucleosome-free regions intermingled with nucleosomes modified with H3K4me3 and H3K27ac, suggesting a dynamic unstable chromatin structure. We further show that human genes involved in cell identity have gene bodies with the same chromatin-instability features as the chicken polychromatic erythrocyte genes participating in the innate immune response.

SARS-CoV-2 multifaceted interaction with the human host. Part II: Innate immunity response, immunopathology, and epigenetics.

The SARS-CoV-2 makes its way into the cell via the ACE2 receptor and the proteolytic action of TMPRSS2. In response to the SARS-CoV-2 infection, the innate immune response is the first line of defense, triggering multiple signaling pathways to produce interferons, pro-inflammatory cytokines and chemokines, and initiating the adaptive immune response against the virus. Unsurprisingly, the virus has developed strategies to evade detection, which can result in delayed, excessive activation of the innate immune system. The response elicited by the host depends on multiple factors, including health status, age, and sex. An overactive innate immune response can lead to a cytokine storm, inflammation, and vascular disruption, leading to the vast array of symptoms exhibited by COVID-19 patients. What is known about the expression and epigenetic regulation of the ACE2 gene and the various players in the host response are explored in this review.

SARS-CoV-2 multifaceted interaction with human host. Part I: What we have learnt and done so far, and the still unknown realities.

SARS-CoV-2, the causing agent of the ongoing COVID-19 pandemic, is a beta-coronavirus which has 80% genetic homology with SARS-CoV, but displays increased virulence and transmissibility. Initially, SARS-CoV-2 was considered a respiratory virus generally causing a mild disease, only severe and fatal in the elderly and individuals with underlying conditions. Severe illnesses and fatalities were attributed to a cytokine storm, an excessive response from the host immune system. However, with the number of infections over 10 millions and still soaring, the insidious and stealthy nature of the virus has emerged, as it causes a vast array of diverse unexpected symptoms among infected individuals, including the young and healthy. It has become evident that besides infecting the respiratory tract, SARS-CoV-2 can affect many organs, possibly through the infection of the endothelium. This review presents an overview of our learning curve with the novel virus emergence, transmission, pathology, biological properties and host-interactions. It also briefly describes remedial measures taken until an effective vaccine is available, that is non-pharmaceutical interventions to reduce the viral spread and the repurposing of existing drugs, approved or in development for other conditions to eliminate the virus or mitigate the cytokine storm.

Global DNA Methylation and Histone Posttranslational Modifications in Human and Nonhuman Primate Brain in Association with Prenatal Alcohol Exposure.

BACKGROUND: Based upon experimental animal studies, the neurodevelopmental abnormalities associated with prenatal alcohol exposure (PNAE)/fetal alcohol spectrum disorder (FASD) have been attributed, at least in part, to epigenetic modifications. However, there are no direct analyses of human brain tissue. METHODS: Immunohistochemical detection of global epigenetic markers was performed on temporal lobe samples of autopsied fetuses and infants with documented PNAE. They were compared to age-, sex-, and postmortem delay-matched control cases (18 pairs; 20 to 70.5 weeks postconception). Temporal lobe tissue from a macaque monkey model of PNAE was also studied (5.7 to 6 months of age). We used antibodies targeting 4 DNA cytosine, 4 histone methylation, and 6 histone acetylation modifications and assigned scores based upon the semiquantitatively graded intensity and proportion of positively labeled nuclei in the ventricular and subventricular zones, ependyma, temporal cortex, temporal white matter, dentate gyrus (DG), and CA1 pyramidal layer. RESULTS: Temporal changes were identified for almost all marks according to the state of maturation in the human brain. In the DG (and 3 other brain regions), a statistically significant increase in H3K9ac was associated with PNAE. Statistically significant decreases were seen among 5mC, H3K4me3, H3K9ac, H3K27ac, H4K12ac, and H4K16ac in select regions. In the macaques, H3K36me3 decreased in the DG, and the ependyma showed decreases in 5fC and H3K36me3. CONCLUSIONS: In human brain, global intranuclear epigenetic modifications are brain region and maturation state-specific. These exploratory results support the general hypothesis that PNAE is associated with a global decrease in DNA methylation, a global decrease in histone methylation, and a global increase in histone acetylation. Although the human and monkey subjects are not directly comparable in terms of brain maturation, considering the rapid temporal changes in global epigenetic modifications during brain development, interspecies comparisons may be extremely difficult.

DNA Methylation Contributes to the Differential Expression Levels of Mecp2 in Male Mice Neurons and Astrocytes.

Methyl CpG binding protein-2 (MeCP2) isoforms (E1 and E2) are important epigenetic regulators in brain cells. Accordingly, MeCP2 loss- or gain-of-function mutation causes neurodevelopmental disorders, including Rett syndrome (RTT), MECP2 duplication syndrome (MDS), and autism spectrum disorders (ASD). Within different types of brain cells, highest MeCP2 levels are detected in neurons and the lowest in astrocytes. However, our current knowledge of Mecp2/MeCP2 regulatory mechanisms remains largely elusive. It appears that there is a sex-dependent effect in X-linked MeCP2-associated disorders, as RTT primarily affects females, whereas MDS is found almost exclusively in males. This suggests that Mecp2 expression levels in brain cells might be sex-dependent. Here, we investigated the sex- and cell type-specific expression of Mecp2 isoforms in male and female primary neurons and astrocytes isolated from the murine forebrain. Previously, we reported that DNA methylation of six Mecp2 regulatory elements correlated with Mecp2 levels in the brain. We now show that in male brain cells, DNA methylation is significantly correlated with the transcript expression of these two isoforms. We show that both Mecp2 isoforms are highly expressed in male neurons compared to male astrocytes, with Mecp2e1 expressed at higher levels than Mecp2e2. Our data indicate that higher DNA methylation at the Mecp2 regulatory element(s) is associated with lower levels of Mecp2 isoforms in male astrocytes compared to male neurons.

Transcription-dependent association of HDAC2 with active chromatin.

Histone deacetylase 2 (HDAC2) catalyzes deacetylation of histones at the promoter and coding regions of transcribed genes and regulates chromatin structure and transcription. To explore the role of HDAC2 and phosphorylated HDAC2 in gene regulation, we studied the location along transcribed genes, the mode of recruitment and the associated proteins with HDAC2 and HDAC2S394ph in chicken polychromatic erythrocytes. We show that HDAC2 and HDAC2S394ph are associated with transcriptionally active chromatin and located in the interchromatin channels. HDAC2S394ph was present primarly at the upstream promoter region of the transcribed CA2 and GAS41 genes, while total HDAC2 was also found within the coding region of the CA2 gene. Recruitment of HDAC2 to these genes was partially dependent upon on-going transcription. Unmodified HDAC2 was associated with RNA binding proteins and interacted with RNA bound to the initiating and elongating forms of RNA polymerase II. HDAC2S394ph was not associated with RNA polymerase II. These results highlight the differential properties of unmodified and phosphorylated HDAC2 and the organization of acetylated transcriptionally active chromatin in the chicken polychromatic erythrocyte.

A 16 Yin Yang gene expression ratio signature for ER+/node- breast cancer.

Breast cancer is one of the leading causes of cancer death in women. It is a complex and heterogeneous disease with different clinical outcomes. Stratifying patients into subgroups with different outcomes could help guide clinical decision making. In this study, we used two opposing groups of genes, Yin and Yang, to develop a prognostic expression ratio signature. Using the METABRIC cohort we identified a16-gene signature capable of stratifying breast cancer patients into four risk levels with intention that low-risk patients would not undergo adjuvant systemic therapy, intermediate-low-risk patients will be treated with hormonal therapy only, and intermediate-high- and high-risk groups will be treated by chemotherapy in addition to the hormonal therapy. The 16-gene signature for four risk level stratifications of breast cancer patients has been validated using 14 independent datasets. Notably, the low-risk group (n = 51) of 205 estrogen receptor-positive and node negative (ER+/node-) patients from three different datasets who had not had any systemic adjuvant therapy had 100% 15-year disease-specific survival rate. The Concordance Index of YMR for ER+/node negative patients is close to the commercially available signatures. However, YMR showed more significance (HR = 3.7, p = 8.7e-12) in stratifying ER+/node- subgroup than OncotypeDx (HR = 2.7, p = 1.3e-7), MammaPrint (HR = 2.5, p = 5.8e-7), rorS (HR = 2.4, p = 1.4e-6), and NPI (HR = 2.6, p = 1.2e-6). YMR signature may be developed as a clinical tool to select a subgroup of low-risk ER+/node- patients who do not require any adjuvant hormonal therapy (AHT).

The discovery and development of the CRISPR system in applications in genome manipulation.

The clustered regularly interspaced short palindromic repeat (CRISPR) associated 9 (Cas9) system is a microbial adaptive immune system that has been recently developed for genomic engineering. From the moment the CRISPR system was discovered in Escherichia coli, the drive to understand the mechanism prevailed, leading to rapid advancement in the knowledge and applications of the CRISPR system. With the ability to characterize and understand the function of the Cas9 endonuclease came the ability to adapt the CRISPR-Cas9 system for use in a variety of applications and disciplines ranging from agriculture to biomedicine. This review will provide a brief overview of the discovery and development of the CRISPR-Cas9 system in applications such as genome regulation and epigenome engineering, as well as the challenges faced.

Dynamic Histone Acetylation of H3K4me3 Nucleosome Regulates MCL1 Pre-mRNA Splicing.

Pre-mRNA splicing is a cotranscriptional process affected by the chromatin architecture along the body of coding genes. Recruited to the pre-mRNA by splicing factors, histone deacetylases (HDACs) and K-acetyltransferases (KATs) catalyze dynamic histone acetylation along the gene. In colon carcinoma HCT 116 cells, HDAC inhibition specifically increased KAT2B occupancy as well as H3 and H4 acetylation of the H3K4 trimethylated (H3K4me3) nucleosome positioned over alternative exon 2 of the MCL1 gene, an event paralleled with the exclusion of exon 2. These results were reproduced in MDA-MB-231, but not in MCF7 breast adenocarcinoma cells. These later cells have much higher levels of demethylase KDM5B than either HCT 116 or MDA-MB-231 cells. We show that H3K4me3 steady-state levels and H3K4me3 occupancy at the end of exon 1 and over exon 2 of the MCL1 gene were lower in MCF7 than in MDA-MB-231 cells. Furthermore, in MCF7 cells, there was minimal effect of HDAC inhibition on H3/H4 acetylation and H3K4me3 levels along the MCL1 gene and no change in pre-mRNA splicing choice. These results show that, upon HDAC inhibition, the H3K4me3 mark plays a critical role in the exclusion of exon 2 from the MCL1 pre-mRNA. J. Cell. Physiol. 231: 2196-2204, 2016. © 2016 Wiley Periodicals, Inc.