Gene induction and repression during terminal erythropoiesis are mediated by distinct epigenetic changes

Abstract It is unclear how epigenetic changes regulate the induction of erythroid-specific genes during terminal erythropoiesis. Here we use global mRNA sequencing (mRNA-seq) and chromatin immunoprecipitation coupled to high-throughput sequencing (CHIP-seq) to investigate the changes that occur in mRNA levels, RNA polymerase II (Pol II) occupancy, and multiple posttranslational histone modifications when erythroid progenitors differentiate into late erythroblasts. Among genes induced during this developmental transition, there was an increase in the occupancy of Pol II, the activation marks H3K4me2, H3K4me3, H3K9Ac, and H4K16Ac, and the elongation methylation mark H3K79me2. In contrast, genes that were repressed during differentiation showed relative decreases in H3K79me2 levels yet had levels of Pol II binding and active histone marks similar to those in erythroid progenitors. We also found that relative changes in histone modification levels, in particular, H3K79me2 and H4K16ac, were most predictive of gene expression patterns. Our results suggest that in terminal erythropoiesis both promoter and elongation-associated marks contribute to the induction of erythroid genes, whereas gene repression is marked by changes in histone modifications mediating Pol II elongation. Our data map the epigenetic landscape of terminal erythropoiesis and suggest that control of transcription elongation regulates gene expression during terminal erythroid differentiation.

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A Dynamic Intron Retention Program in the Mammalian Megakaryocyte and Erythrocyte Lineages

Blood ◽

10.1182/blood.v126.23.2380.2380 ◽

2015 ◽

Vol 126 (23) ◽

pp. 2380-2380

Author(s):

Christopher R Edwards ◽

Rob Middleton ◽

Xiuli An ◽

Tejaswini Mishra ◽

Narla Mohandas ◽

...

Keyword(s):

Gene Expression ◽

Alternative Splicing ◽

Conflicts Of Interest ◽

Intron Retention ◽

Expression Patterns ◽

Erythroid Differentiation ◽

Cell Types ◽

Mrna Levels ◽

Protein Coding ◽

Nonsense Mediated Decay

Abstract Intron retention (IR), the least studied form of alternative splicing, has recently been shown to have important biological roles in a variety of cell types. While it can alter a gene's protein-coding sequence, it is becoming particularly well-known for its potential to impact gene expression by destabilizing mRNAs through the nonsense-mediated decay pathway or by promoting their retention in the nucleus. A complex, dynamic, and biologically important IR program has been described in maturing mammalian granulocytes, but it is unknown whether IR occurs broadly in other hematopoietic lineages. We therefore globally assessed IR in the mammalian erythroid and megakaryocyte lineages. Intron Retention Finder, a bioinformatics tool that measures IR in RNA-seq datasets, was used to analyze IR in primary cells of the erythroid and megakaryocyte lineages as well as their common progenitor cells. Both lineages exhibit an extensive differential IR program involving hundreds of introns and genes. Complex IR patterns were seen in murine erythropoiesis from the megakaryocytic-erythroid branch point throughout the terminal maturation stages. Within the terminally differentiating proerythroblast to orthochromatic erythroblast stages, hundreds of introns saw their retention level increase as cells differentiate while a smaller set exhibited an opposing trend. Similarly complex patterns including a dramatic IR increase in orthochromatic erythroblasts were observed during human terminal erythroid differentiation, but not involving the murine orthologous introns or genes. Despite the common origin of erythroid cells and megakaryocytes and their overlapping gene expression patterns, the megakaryocytic IR program is entirely distinct from that of the erythroid lineage with regards to introns, genes, and affected gene ontologies. This suggests that the dynamic IR patterns are not simply the result of general maturational changes, but rather may arise via lineage-specific mechanisms. Importantly, we observed an inverse relationship between IR and gene expression changes, supporting the hypothesis that IR serves to regulate mRNA levels. Our findings add a new dimension to the megakaryocyte and erythroid transcription programs by expanding the mechanisms of gene control to include this understudied form of alternative splicing. Disclosures No relevant conflicts of interest to declare.

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Histone modifications form a cell-type-specific chromosomal bar code that persists through the cell cycle

Scientific Reports ◽

10.1038/s41598-021-82539-z ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

John A. Halsall ◽

Simon Andrews ◽

Felix Krueger ◽

Charlotte E. Rutledge ◽

Gabriella Ficz ◽

...

Keyword(s):

Gene Expression ◽

Cell Cycle ◽

Histone Modifications ◽

Expression Patterns ◽

Cell Types ◽

Cell Type ◽

Bar Code ◽

Genes Encoding ◽

Cell Type Specific ◽

Rolling Windows

AbstractChromatin configuration influences gene expression in eukaryotes at multiple levels, from individual nucleosomes to chromatin domains several Mb long. Post-translational modifications (PTM) of core histones seem to be involved in chromatin structural transitions, but how remains unclear. To explore this, we used ChIP-seq and two cell types, HeLa and lymphoblastoid (LCL), to define how changes in chromatin packaging through the cell cycle influence the distributions of three transcription-associated histone modifications, H3K9ac, H3K4me3 and H3K27me3. We show that chromosome regions (bands) of 10–50 Mb, detectable by immunofluorescence microscopy of metaphase (M) chromosomes, are also present in G1 and G2. They comprise 1–5 Mb sub-bands that differ between HeLa and LCL but remain consistent through the cell cycle. The same sub-bands are defined by H3K9ac and H3K4me3, while H3K27me3 spreads more widely. We found little change between cell cycle phases, whether compared by 5 Kb rolling windows or when analysis was restricted to functional elements such as transcription start sites and topologically associating domains. Only a small number of genes showed cell-cycle related changes: at genes encoding proteins involved in mitosis, H3K9 became highly acetylated in G2M, possibly because of ongoing transcription. In conclusion, modified histone isoforms H3K9ac, H3K4me3 and H3K27me3 exhibit a characteristic genomic distribution at resolutions of 1 Mb and below that differs between HeLa and lymphoblastoid cells but remains remarkably consistent through the cell cycle. We suggest that this cell-type-specific chromosomal bar-code is part of a homeostatic mechanism by which cells retain their characteristic gene expression patterns, and hence their identity, through multiple mitoses.

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Gene expression profiling of epigenetic chromatin modification enzymes and histone marks by cigarette smoke: implications for COPD and lung cancer

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00253.2016 ◽

2016 ◽

Vol 311 (6) ◽

pp. L1245-L1258 ◽

Cited By ~ 19

Author(s):

Isaac K. Sundar ◽

Irfan Rahman

Keyword(s):

Gene Expression ◽

Lung Cancer ◽

Dna Methylation ◽

Cigarette Smoke ◽

Histone Modifications ◽

Expression Patterns ◽

Chromatin Modification ◽

Gene Expression Patterns ◽

Validation Data ◽

Histone Marks

Chromatin-modifying enzymes mediate DNA methylation and histone modifications on recruitment to specific target gene loci in response to various stimuli. The key enzymes that regulate chromatin accessibility for maintenance of modifications in DNA and histones, and for modulation of gene expression patterns in response to cigarette smoke (CS), are not known. We hypothesize that CS exposure alters the gene expression patterns of chromatin-modifying enzymes, which then affects multiple downstream pathways involved in the response to CS. We have, therefore, analyzed chromatin-modifying enzyme profiles and validated by quantitative real-time PCR (qPCR). We also performed immunoblot analysis of targeted histone marks in C57BL/6J mice exposed to acute and subchronic CS, and of lungs from nonsmokers, smokers, and patients with chronic obstructive pulmonary disease (COPD). We found a significant increase in expression of several chromatin modification enzymes, including DNA methyltransferases, histone acetyltransferases, histone methyltransferases, and SET domain proteins, histone kinases, and ubiquitinases. Our qPCR validation data revealed a significant downregulation of Dnmt1, Dnmt3a, Dnmt3b, Hdac2, Hdac4, Hat1, Prmt1, and Aurkb. We identified targeted chromatin histone marks (H3K56ac and H4K12ac), which are induced by CS. Thus CS-induced genotoxic stress differentially affects the expression of epigenetic modulators that regulate transcription of target genes via DNA methylation and site-specific histone modifications. This may have implications in devising epigenetic-based therapies for COPD and lung cancer.

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Gene expression analysis of the pro-oestrous-stage rat uterus reveals neuroligin 2 as a novel steroid-regulated gene

Reproduction Fertility and Development ◽

10.1071/rd04040 ◽

2004 ◽

Vol 16 (8) ◽

pp. 763 ◽

Cited By ~ 8

Author(s):

Han-Seung Kang ◽

Chae-Kwan Lee ◽

Ju-Ran Kim ◽

Seong-Jin Yu ◽

Sung-Goo Kang ◽

...

Keyword(s):

Gene Expression ◽

Northern Blot ◽

Blot Analysis ◽

Northern Blot Analysis ◽

Expression Profiles ◽

Expression Patterns ◽

Oestrous Cycle ◽

Mrna Levels ◽

Rat Uterus ◽

Ovx Rats

In the present study, differential gene expression in the uteri of ovariectomised (OVX) and pro-oestrous rats (OVX v. pro-oestrus pair) was investigated using cDNA expression array analysis. Differential uterine gene expression in OVX rats and progesterone (P4)-injected OVX rats (OVX v. OVX + P4 pair) was also examined. The uterine gene expression profiles of these two sets of animals were also compared for the effects of P4 treatment. RNA samples were extracted from uterine tissues and reverse transcribed in the presence of [α32P]-dATP. Membrane sets of rat arrays were hybridised with cDNA probe sets. Northern blot analysis was used to validate the relative gene expression patterns obtained from the cDNA array. Of the 1176 cDNAs examined, 23 genes showed significant (>two-fold) changes in expression in the OVX v. pro-oestrus pair. Twenty of these genes were upregulated during pro-oestrus compared with their expression in the OVX rat uterus. In the OVX v. OVX + P4 pair, 22 genes showed significant (>two-fold) changes in gene expression. Twenty of these genes were upregulated in the OVX + P4 animals. The genes for nuclear factor I–XI, afadin, neuroligin 2, semaphorin Z, calpain 4, cyclase-associated protein homologue, thymosin β-4X and p8 were significantly upregulated in the uteri of the pro-oestrus and OVX + P4 rats of both experimental pairs compared with the OVX rat uteri. These genes appear to be under the control of P4. One of the most interesting findings of the present study is the unexpected and marked expression of the neuroligin 2 gene in the rat uterus. This gene is expressed at high levels in the central nervous system and acts as a nerve cell adhesion factor. According to Northern blot analysis, neuroligin 2 gene expression was higher during the pro-oestrus and metoestrus stages than during the oestrus and dioestrus stages of the oestrous cycle. In addition, neuroligin 2 mRNA levels were increased by both 17β-oestradiol (E2) and P4, although P4 administration upregulated gene expression to a greater extent than injection of E2. These results indicate that neuroligin 2 gene expression in the rat uterus is under the control of both E2 and P4, which are secreted periodically during the oestrous cycle.

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Independent Recruitment of Mediator and SAGA by the Activator Met4

Molecular and Cellular Biology ◽

10.1128/mcb.26.8.3149-3163.2006 ◽

2006 ◽

Vol 26 (8) ◽

pp. 3149-3163 ◽

Cited By ~ 28

Author(s):

Christophe Leroy ◽

Laëtitia Cormier ◽

Laurent Kuras

Keyword(s):

Gene Expression ◽

Amino Acids ◽

Rna Polymerase Ii ◽

Gene Network ◽

Pol Ii ◽

Eukaryotic Gene Expression ◽

Eukaryotic Gene ◽

Target Promoters ◽

Sulfur Containing

ABSTRACT Mediator is a key RNA polymerase II (Pol II) cofactor in the regulation of eukaryotic gene expression. It is believed to function as a coactivator linking gene-specific activators to the basal Pol II initiation machinery. In support of this model, we provide evidence that Mediator serves in vivo as a coactivator for the yeast activator Met4, which controls the gene network responsible for the biosynthesis of sulfur-containing amino acids and S-adenosylmethionine. In addition, we show that SAGA (Spt-Ada-Gcn5-acetyltransferase) is also recruited to Met4 target promoters, where it participates in the recruitment of Pol II by a mechanism involving histone acetylation. Interestingly, we find that SAGA is not required for Mediator recruitment by Met4 and vice versa. Our results provide a novel example of functional interplay between Mediator and coactivators involved in histone modification.

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A machine learning-based framework for modeling transcription elongation

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2007450118 ◽

2021 ◽

Vol 118 (6) ◽

pp. e2007450118

Author(s):

Peiyuan Feng ◽

An Xiao ◽

Meng Fang ◽

Fangping Wan ◽

Shuya Li ◽

...

Keyword(s):

Rna Polymerase Ii ◽

High Throughput Sequencing ◽

Gene Expression Regulation ◽

Transcription Elongation ◽

Transcriptional Elongation ◽

Sequencing Data ◽

Pol Ii ◽

High Throughput Sequencing Data ◽

Pol Ii Pausing ◽

Elongation Process

RNA polymerase II (Pol II) generally pauses at certain positions along gene bodies, thereby interrupting the transcription elongation process, which is often coupled with various important biological functions, such as precursor mRNA splicing and gene expression regulation. Characterizing the transcriptional elongation dynamics can thus help us understand many essential biological processes in eukaryotic cells. However, experimentally measuring Pol II elongation rates is generally time and resource consuming. We developed PEPMAN (polymerase II elongation pausing modeling through attention-based deep neural network), a deep learning-based model that accurately predicts Pol II pausing sites based on the native elongating transcript sequencing (NET-seq) data. Through fully taking advantage of the attention mechanism, PEPMAN is able to decipher important sequence features underlying Pol II pausing. More importantly, we demonstrated that the analyses of the PEPMAN-predicted results around various types of alternative splicing sites can provide useful clues into understanding the cotranscriptional splicing events. In addition, associating the PEPMAN prediction results with different epigenetic features can help reveal important factors related to the transcription elongation process. All these results demonstrated that PEPMAN can provide a useful and effective tool for modeling transcription elongation and understanding the related biological factors from available high-throughput sequencing data.

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GDF-9 and BMP-15 mRNA Levels in Canine Cumulus Cells Related to Cumulus Expansion and the Maturation Process

Animals ◽

10.3390/ani10030462 ◽

2020 ◽

Vol 10 (3) ◽

pp. 462 ◽

Cited By ~ 3

Author(s):

George Ramirez ◽

Jaime Palomino ◽

Karla Aspee ◽

Monica De los Reyes

Keyword(s):

Gene Expression ◽

Estrous Cycle ◽

Expression Patterns ◽

Cumulus Cells ◽

Mrna Levels ◽

Cumulus Expansion ◽

Polymerase Chain ◽

Specific Regulation ◽

Reproductive Phases

The competence to undergo expansion is a characteristic of cumulus cells (CCs). The aim was to investigate the expression of GDF-9 and BMP-15 mRNA in canine cumulus cells in relation to cumulus expansion and meiotic development over the estrous cycle. CCs were recovered from nonmatured and in vitro-matured (IVM) dog cumulus oocyte complexes (COCs), which were obtained from antral follicles at different phases of the estrous cycle. Quantitative real-time polymerase chain reaction (q-PCR) was used to evaluate the relative abundance of GDF-9 and BMP-15 transcripts from the CCs with or without signs of expansion. The results were evaluated by ANOVA and logistic regression. The maturity of the oocyte and the expansion process affected the mRNA levels in CCs. There were differences (p < 0.05) in GDF-9 and BMP-15 gene expression in CCs isolated from nonmatured COCs when comparing the reproductive phases. Lower mRNA levels (p < 0.05) were observed in anestrus and proestrus in comparison to those in estrus and diestrus. In contrast, when comparing GDF-9 mRNA levels in IVM COCs, no differences were found among the phases of the estrous cycle in expanded and nonexpanded CCs (p < 0.05). However, the highest (p < 0.05) BMP-15 gene expression in CCs that did not undergo expansion was exhibited in anestrus and the lowest (p < 0.05) expression was observed in estrus in expanded CCs. Although the stage of the estrous cycle did not affect the second metaphase (MII )rates, the expanded CCs obtained at estrus coexisted with higher percentages of MII (p < 0.05). In conclusion, the differential expression patterns of GDF-9 and BMP-15 mRNA transcripts might be related to cumulus expansion and maturation processes, suggesting specific regulation and temporal changes in their expression.

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Gene Expression Patterns Distinguish Mortality Risk in Patients with Postsurgical Shock

Journal of Clinical Medicine ◽

10.3390/jcm9051276 ◽

2020 ◽

Vol 9 (5) ◽

pp. 1276

Author(s):

Pedro Martínez-Paz ◽

Marta Aragón-Camino ◽

Esther Gómez-Sánchez ◽

Mario Lorenzo-López ◽

Estefanía Gómez-Pesquera ◽

...

Keyword(s):

Gene Expression ◽

High Risk ◽

Mortality Risk ◽

Validation Cohort ◽

Expression Patterns ◽

Gene Expression Signature ◽

Mrna Levels ◽

Characteristic Analysis ◽

Discovery Cohort ◽

Risk Of Death

Nowadays, mortality rates in intensive care units are the highest of all hospital units. However, there is not a reliable prognostic system to predict the likelihood of death in patients with postsurgical shock. Thus, the aim of the present work is to obtain a gene expression signature to distinguish the low and high risk of death in postsurgical shock patients. In this sense, mRNA levels were evaluated by microarray on a discovery cohort to select the most differentially expressed genes between surviving and non-surviving groups 30 days after the operation. Selected genes were evaluated by quantitative real-time polymerase chain reaction (qPCR) in a validation cohort to validate the reliability of data. A receiver-operating characteristic analysis with the area under the curve was performed to quantify the sensitivity and specificity for gene expression levels, which were compared with predictions by established risk scales, such as acute physiology and chronic health evaluation (APACHE) and sequential organ failure assessment (SOFA). IL1R2, CD177, RETN, and OLFM4 genes were upregulated in the non-surviving group of the discovery cohort, and their predictive power was confirmed in the validation cohort. This work offers new biomarkers based on transcriptional patterns to classify the postsurgical shock patients according to low and high risk of death. The results present more accuracy than other mortality risk scores.

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Expression of Inflammation-Related Genes Is Altered in Gastric Tissue of Patients with Advanced Stages of NAFLD

Mediators of Inflammation ◽

10.1155/2013/684237 ◽

2013 ◽

Vol 2013 ◽

pp. 1-10 ◽

Cited By ~ 5

Author(s):

Rohini Mehta ◽

Aybike Birerdinc ◽

Arpan Neupane ◽

Amirhossein Shamsaddini ◽

Arian Afendy ◽

...

Keyword(s):

Gene Expression ◽

Hepatic Fibrosis ◽

Interleukin 1 ◽

Expression Patterns ◽

Interleukin 8 ◽

Morbidly Obese ◽

Low Grade ◽

Mrna Levels ◽

Gastric Tissue ◽

Hepatic Inflammation

Obesity is associated with chronic low-grade inflammation perpetuated by visceral adipose. Other organs, particularly stomach and intestine, may also overproduce proinflammatory molecules. We examined the gene expression patterns in gastric tissue of morbidly obese patients with nonalcoholic fatty liver disease (NAFLD) and compared the changes in gene expression in different histological forms of NAFLD. Stomach tissue samples from 20 morbidly obese NAFLD patients who were undergoing sleeve gastrectomy were profiled using qPCR for 84 genes encoding inflammatory cytokines, chemokines, their receptors, and other components of inflammatory cascades. Interleukin 8 receptor-beta (IL8RB) gene overexpression in gastric tissue was correlated with the presence of hepatic steatosis, hepatic fibrosis, and histologic diagnosis of nonalcoholic steatohepatitis (NASH). Expression levels of soluble interleukin 1 receptor antagonist (IL1RN) were correlated with the presence of NASH and hepatic fibrosis. mRNA levels of interleukin 8 (IL8), chemokine (C-C motif) ligand 4 (CCL4), and its receptor chemokine (C-C motif) receptor type 5 (CCR5) showed a significant increase in patients with advanced hepatic inflammation and were correlated with the severity of the hepatic inflammation. The results of our study suggest that changes in expression patterns for inflammatory molecule encoding genes within gastric tissue may contribute to the pathogenesis of obesity-related NAFLD.

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Lenalidomide and Novel Immunomodulatory Drugs (IMiDs®): A New Approach to the Regulation of Erythropoiesis and Hemoglobin Synthesis in Anemia.

Blood ◽

10.1182/blood.v106.11.2702.2702 ◽

2005 ◽

Vol 106 (11) ◽

pp. 2702-2702 ◽

Cited By ~ 2

Author(s):

Laure Moutouh de Parseval ◽

Helen Brady ◽

Dominique Verhelle ◽

Laura G. Corral ◽

Emilia Glezer ◽

...

Keyword(s):

Gene Expression ◽

Globin Gene ◽

Fetal Hemoglobin ◽

Erythroid Differentiation ◽

Chemotherapeutic Agents ◽

Immunomodulatory Drugs ◽

Erythroid Cells ◽

Erythroid Progenitors ◽

Hemoglobin Synthesis ◽

Hematological Cancers

Abstract Clinical trial results have demonstrated that lenalidomide (Revlimid®) reduces or even eliminates the need for red blood cell transfusions in some anemic myelodysplastic patients. We have examined whether lenalidomide and Actimid™, members of a new class of immunomodulatory drugs (IMiDs®), which are currently under evaluation for the treatment of hematological cancers could regulate erythropoiesis and hemoglobin synthesis. For this purpose, we used an in vitro culture model to differentiate human erythroid progenitors from bone marrow or peripheral blood CD34+ cells. We demonstrate that lenalidomide and AztimidTM modulate erythropoiesis and increase proliferation of immature erythroid cells. In addition to the regulation of erythroid differentiation, lenalidomide and ActimidTM are potent inducers of fetal hemoglobin. Unlike other inducers of fetal hemoglobin such as 5-aza-cytidine that are cytotoxic, IMiDs® promoted survival of erythroblast cultured with known cytotoxic drug. Gene expression profiling of erythroid differentiated cells showed that IMiDs® regulate specific erythroid transcription factors and genes that participate in hemoglobin synthesis, and genes invoved in cell cycle and cellular differentiation. Globin gene expression is controlled by IMiDs® during erythroid differentiation by inducing fetal hemoglobin synthesis. Our results support the hypothesis that IMiDs® restore effective erythropoiesis in myelodysplastic patients and protect erythroid cells from the cytotoxic effect of chemotherapeutic agents. In conclusion, IMiDs® may represent an interesting new therapy for cancer-related anemia and β-hemoglobinopathies.

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