scholarly journals Histone H3 Lysine 4 methyltransferases MLL3 and MLL4 Modulate Long-range Chromatin Interactions at Enhancers

2017 ◽  
Author(s):  
Jian Yan ◽  
Shi-An A Chen ◽  
Andrea Local ◽  
Tristin Liu ◽  
Yunjiang Qiu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Long Range ◽  
Mammalian Cells ◽  
Regulation Of Gene Expression ◽  
Histone H3 ◽  
Embryonic Stem ◽  
Active Role ◽  
Specific Gene ◽  
Chromatin Interactions

SUMMARYRegulation of gene expression in mammalian cells depends on long-range chromatin interactions between enhancers and promoters. Currently, the exact mechanisms that connect distal enhancers to their specific target promoters remain to be fully elucidated. Here we show that the histone H3 Lysine 4 monomethylation (H3K4me1) writer proteins MLL3 and MLL4 (MLL3/4) play an active role in this process. We demonstrate that in differentiating mouse embryonic stem cells, MLL3/4-dependent deposition of H3K4me1 at enhancers correlates with increased levels of chromatin interactions, whereas loss of MLL3/4 leads to greatly reduced frequencies of chromatin interactions and failure of lineage-specific gene expression programs. We further show that H3K4me1 facilitates recruitment of the Cohesin complex to chromatin in vitro and in vivo, providing a potential mechanism for MLL3/4 to promote chromatin looping. Taken together, our results support an active role for MLL3/4 in modulating chromatin organization at enhancers in mammalian cells.

scholarly journals The TAZ2 domain of CBP/p300 directs acetylation towards H3K27 within chromatin

2020 ◽  
Author(s):  
Thomas W. Sheahan ◽  
Viktoria Major ◽  
Kimberly M. Webb ◽  
Elana Bryan ◽  
Philipp Voigt
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Enzymatic Activity ◽  
Crucial Role ◽  
Regulation Of Gene Expression ◽  
Histone H3 ◽  
Embryonic Stem ◽  
Site Specific ◽  
Lysine Residues

AbstractThe closely related acetyltransferases CBP and p300 are key regulators of gene expression in metazoans. CBP/p300 acetylate several specific lysine residues within nucleosomes, including histone H3 lysine 27 (H3K27), a hallmark of active enhancers and promoters. However, it has remained largely unclear how specificity of CBP/p300 towards H3K27 is achieved. Here we show that the TAZ2 domain of CBP is required for efficient acetylation of H3K27, while curbing activity towards other lysine residues within nucleosomes. We find that TAZ2 is a sequence-independent DNA binding module, promoting interaction between CBP and nucleosomes, thereby enhancing enzymatic activity and regulating substrate specificity of CBP. TAZ2 is further required to stabilize CBP binding to chromatin in mouse embryonic stem cells, facilitating specificity towards H3K27 and modulating gene expression. These findings reveal a crucial role of TAZ2 in regulating H3K27ac, while highlighting the importance of correct site-specific acetylation for proper regulation of gene expression.

Murine gastrulation requires HNF-4 regulated gene expression in the visceral endoderm: tetraploid rescue of Hnf-4(−/−) embryos

Development ◽  
1997 ◽  
Vol 124 (2) ◽  
pp. 279-287 ◽  
Author(s):  
S.A. Duncan ◽  
A. Nagy ◽  
W. Chan
Keyword(s):  
Gene Expression ◽  
Genetic Manipulation ◽  
Es Cells ◽  
Critical Role ◽  
Embryonic Stem ◽  
Retinol Binding Protein ◽  
Specific Gene ◽  
Visceral Endoderm ◽  
Regulated Gene Expression

Immediately prior to gastrulation the murine embryo consists of an outer layer of visceral endoderm (VE) and an inner layer of ectoderm. Differentiation and migration of the ectoderm then occurs to produce the three germ layers (ectoderm, embryonic endoderm and mesoderm) from which the fetus is derived. An indication that the VE might have a critical role in this process emerged from studies of Hnf-4(−/−) mouse embryos which fail to undergo normal gastrulation. Since expression of the transcription factor HNF-4 is restricted to the VE during this phase of development, we proposed that HNF-4-regulated gene expression in the VE creates an environment capable of supporting gastrulation. To address this directly we have exploited the versatility of embryonic stem (ES) cells which are amenable to genetic manipulation and can be induced to form VE in vitro. Moreover, embryos derived solely from ES cells can be generated by aggregation with tetraploid morulae. Using Hnf-4(−/−) ES cells we demonstrate that HNF-4 is a key regulator of tissue-specific gene expression in the VE, required for normal expression of secreted factors including alphafetoprotein, apolipoproteins, transthyretin, retinol binding protein, and transferrin. Furthermore, specific complementation of Hnf-4(−/−) embryos with tetraploid-derived Hnf-4(+/+) VE rescues their early developmental arrest, showing conclusively that a functional VE is mandatory for gastrulation.

Characterization of hematopoietic lineage-specific gene expression by ES cell in vitro differentiation induction system

Blood ◽  
2000 ◽  
Vol 95 (3) ◽  
pp. 870-878 ◽  
Author(s):  
Takumi Era ◽  
Toshiaki Takagi ◽  
Tomomi Takahashi ◽  
Jean-Christophe Bories ◽  
Toru Nakano
Keyword(s):  
Gene Expression ◽  
Cell Lines ◽  
Embryonic Stem ◽  
Experimental System ◽  
Specific Gene ◽  
Specific Gene Expression ◽  
Differentiation Induction ◽  
Lineage Specific Gene ◽  
Megakaryocytic Cell

The continuous generation of mature blood cells from hematopoietic progenitor cells requires a highly complex series of molecular events. To examine lineage-specific gene expression during the differentiation process, we developed a novel method combiningLacZ reporter gene analysis with in vitro hematopoietic differentiation induction from mouse embryonic stem cells. For a model system using this method, we chose the erythroid and megakaryocytic differentiation pathways. Although erythroid and megakaryocytic cells possess distinct functional and morphologic features, these 2 lineages originate from bipotential erythro-megakaryocytic progenitors and share common lineage-restricted transcription factors. A portion of the 5′ flanking region of the human glycoprotein IIb (IIb) integrin gene extending from base −598 to base +33 was examined in detail. As reported previously, this region is sufficient for megakaryocyte-specific gene expression. However, previous reports that used human erythro-megakaryocytic cell lines suggested that one or more negative regulatory regions were necessary for megakaryocyte-specific gene expression. Our data clearly showed that an approximately 200-base enhancer region extending from −598 to −400 was sufficient for megakaryocyte-specific gene expression. This experimental system has advantages over those using erythro-megakaryocytic cell lines because it recapitulates normal hematopoietic cell development and differentiation. Furthermore, this system is more efficient than transgenic analysis and can easily examine gene expression with null mutations of specific genes.

scholarly journals Small Interfering RNA–Mediated Suppression of Fas Modulate Apoptosis and Proliferation in Rat Intervertebral Disc Cells

2017 ◽  
Vol 11 (5) ◽  
pp. 686-693
Author(s):  
Jong-Beom Park ◽  
Chanjoo Park
Keyword(s):  
Gene Expression ◽  
Mammalian Cells ◽  
Small Interfering Rna ◽  
Mrna Level ◽  
Serum Deprivation ◽  
Specific Gene ◽  
Disc Cells ◽  
Fetal Bovine ◽  
Interfering Rna

<sec><title>Study Design</title><p><italic>In vitro</italic> cell culture model.</p></sec><sec><title>Purpose</title><p>To investigate the effect of small interfering RNA (siRNA) on Fas expression, apoptosis, and proliferation in serum-deprived rat disc cells.</p></sec><sec><title>Overview of Literature</title><p>Synthetic siRNA can trigger an RNA interference (RNAi) response in mammalian cells and precipitate the inhibition of specific gene expression. However, the potential utility of siRNA technology in downregulation of specific genes associated with disc cell apoptosis remains unclear.</p></sec><sec><title>Methods</title><p>Rat disc cells were isolated and cultured in the presence of either 10% fetal bovine serum (FBS) (normal control) or 0% FBS (serum deprivation to induce apoptosis) for 48 hours. Fas expression, apoptosis, and proliferation were determined. Additionally, siRNA oligonucleotides against Fas (Fas siRNA) were transfected into rat disc cells to suppress Fas expression. Changes in Fas expression were assessed by reverse transcription-polymerase chain reaction and semiquantitatively analyzed using densitometry. The effect of Fas siRNA on apoptosis and proliferation of rat disc cells were also determined. Negative siRNA and transfection agent alone (Mock) were used as controls.</p></sec><sec><title>Results</title><p>Serum deprivation increased apoptosis by 40.3% (<italic>p</italic>&lt;0.001), decreased proliferation by 45.3% (<italic>p</italic>&lt;0.001), and upregulated Fas expression. Additionally, Fas siRNA suppressed Fas expression in serum-deprived cultures, with 68.5% reduction at the mRNA level compared to the control cultures (<italic>p</italic>&lt;0.001). Finally, Fas siRNA–mediated suppression of Fas expression significantly inhibited apoptosis by 9.3% and increased proliferation by 21% in serum-deprived cultures (<italic>p</italic>&lt;0.05 for both).</p></sec><sec><title>Conclusions</title><p>The observed dual positive effect of Fas siRNA might be a powerful therapeutic approach for disc degeneration by suppression of harmful gene expression.</p></sec>

scholarly journals Precise tuning of gene expression output levels in mammalian cells

2018 ◽  
Author(s):  
Yale S. Michaels ◽  
Mike B. Barnkob ◽  
Hector Barbosa ◽  
Toni A. Baeumler ◽  
Mary K. Thompson ◽  
...  
Keyword(s):  
Gene Expression ◽  
Mammalian Cells ◽  
High Throughput Sequencing ◽  
Regulation Of Gene Expression ◽  
Antigen Expression ◽  
Control Of Gene Expression ◽  
Precise Control ◽  
Mouse Melanoma Model

ABSTRACTPrecise, analogue regulation of gene expression is critical for development, homeostasis and regeneration in mammals. In contrast, widely employed experimental and therapeutic approaches such as knock-in/out strategies are more suitable for binary control of gene activity, while RNA interference (RNAi) can lead to pervasive off-target effects and unpredictable levels of repression. Here we report on a method for the precise control of gene expression levels in mammalian cells based on engineered, synthetic microRNA response elements (MREs). To develop this system, we established a high-throughput sequencing approach for measuring the efficacy of thousands of miR-17 MRE variants. This allowed us to create a library of microRNA silencing-mediated fine-tuners (miSFITs) of varying strength that can be employed to control the expression of user specified genes. To demonstrate the value of this technology, we used a panel of miSFITs to tune the expression of a peptide antigen in a mouse melanoma model. This analysis revealed that antigen expression level is a key determinant of the anti-tumour immune response in vitro and in vivo. miSFITs are a powerful tool for modulating gene expression output levels with applications in research and cellular engineering.

scholarly journals Functional evaluation of transposable elements as transcriptional enhancers in mouse embryonic and trophoblast stem cells

2018 ◽  
Author(s):  
Christopher D. Todd ◽  
Özgen Deniz ◽  
Miguel R. Branco
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Gene Regulation ◽  
Transposable Elements ◽  
Regulation Of Gene Expression ◽  
Embryonic Stem ◽  
Specific Gene ◽  
Functional Evaluation ◽  
Trophoblast Stem Cells ◽  
Trophoblast Stem

AbstractThe recurrent invasion and expansion of transposable elements (TEs) throughout evolution brought with it a vast array of coding and non-coding sequences that can serve as substrates for natural selection. Namely, TEs are thought to have contributed to the establishment of gene regulatory networks via their cis-acting elements. Both the embryonic and extraembryonic lineages of the early mouse embryo are thought to have benefited from the co-option of TEs as distal enhancer elements. However, there is little to no evidence that these particular TEs play significant roles in the regulation of gene expression. Here we tested for roles of TEs as enhancers in mouse embryonic and trophoblast stem cells by combining bioinformatic analyses with genetic and epigenetic editing experiments. Epigenomic and transcriptomic data from wildtype cells suggested that a large number of TEs played a role in the establishment of highly tissue-specific gene expression programmes. Through genetic editing of individual TEs we confirmed a subset of these regulatory relationships. However, a wider survey via CRISPR interference of RLTR13D6 elements in embryonic stem cells revealed that only a minority play significant roles in gene regulation. Our results suggest that a small proportion of TEs contribute to the mouse pluripotency regulatory network, and highlight the importance of functional experiments when evaluating the role of TEs in gene regulation.

scholarly journals RALYL increases hepatocellular carcinoma stemness by sustaining the mRNA stability of TGF-β2

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Xia Wang ◽  
Jin Wang ◽  
Yu-Man Tsui ◽  
Chaoran Shi ◽  
Ying Wang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mrna Stability ◽  
Rna Binding ◽  
Embryonic Stem ◽  
Molecular Characteristics ◽  
Specific Gene ◽  
Functional Studies ◽  
Poor Differentiation ◽  
Development In Vitro

AbstractGrowing evidences suggest that cancer stem cells exhibit many molecular characteristics and phenotypes similar to their ancestral progenitor cells. In the present study, human embryonic stem cells are induced to differentiate into hepatocytes along hepatic lineages to mimic liver development in vitro. A liver progenitor specific gene, RALY RNA binding protein like (RALYL), is identified. RALYL expression is associated with poor prognosis, poor differentiation, and metastasis in clinical HCC patients. Functional studies reveal that RALYL could promote HCC tumorigenicity, self-renewal, chemoresistance, and metastasis. Moreover, molecular mechanism studies show that RALYL could upregulate TGF-β2 mRNA stability by decreasing N6-methyladenosine (m6A) modification. TGF-β signaling and the subsequent PI3K/AKT and STAT3 pathways, upregulated by RALYL, contribute to the enhancement of HCC stemness. Collectively, RALYL is a liver progenitor specific gene and regulates HCC stemness by sustaining TGF-β2 mRNA stability. These findings may inspire precise therapeutic strategies for HCC.

scholarly journals The dyskerin ribonucleoprotein complex as an OCT4/SOX2 coactivator in embryonic stem cells

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Yick W Fong ◽  
Jaclyn J Ho ◽  
Carla Inouye ◽  
Robert Tjian
Keyword(s):  
Stem Cell ◽  
Es Cells ◽  
Embryonic Stem ◽  
In Vitro Transcription ◽  
Specific Gene ◽  
Transcriptional Level ◽  
Ips Cell ◽  
Ribonucleoprotein Complex ◽  
Transcription System

Acquisition of pluripotency is driven largely at the transcriptional level by activators OCT4, SOX2, and NANOG that must in turn cooperate with diverse coactivators to execute stem cell-specific gene expression programs. Using a biochemically defined in vitro transcription system that mediates OCT4/SOX2 and coactivator-dependent transcription of the Nanog gene, we report the purification and identification of the dyskerin (DKC1) ribonucleoprotein complex as an OCT4/SOX2 coactivator whose activity appears to be modulated by a subset of associated small nucleolar RNAs (snoRNAs). The DKC1 complex occupies enhancers and regulates the expression of key pluripotency genes critical for self-renewal in embryonic stem (ES) cells. Depletion of DKC1 in fibroblasts significantly decreased the efficiency of induced pluripotent stem (iPS) cell generation. This study thus reveals an unanticipated transcriptional role of the DKC1 complex in stem cell maintenance and somatic cell reprogramming.

scholarly journals ARID2 Chromatin Remodeler in Hepatocellular Carcinoma

Cells ◽  
2020 ◽  
Vol 9 (10) ◽  
pp. 2152
Author(s):  
Robin Loesch ◽  
Linda Chenane ◽  
Sabine Colnot
Keyword(s):  
Gene Expression ◽  
Hepatocellular Carcinoma ◽  
Associated Factors ◽  
Regulation Of Gene Expression ◽  
Specific Gene ◽  
Chromatin Remodeler ◽  
Chromatin Remodelers ◽  
Genes Encoding ◽  
Gene Alterations

Chromatin remodelers are found highly mutated in cancer including hepatocellular carcinoma. These mutations frequently occur in ARID (AT-rich Interactive Domain) genes, encoding subunits of the ATP-dependent SWI/SNF remodelers. The increasingly prevalent complexity that surrounds the functions and specificities of the highly modular BAF (BG1/BRM-associated factors) and PBAF (polybromo-associated BAF) complexes, including ARID1A/B or ARID2, is baffling. The involvement of the SWI/SNF complexes in diverse tissues and processes, and especially in the regulation of gene expression, multiplies the specific outcomes of specific gene alterations. A better understanding of the molecular consequences of specific mutations impairing chromatin remodelers is needed. In this review, we summarize what we know about the tumor-modulating properties of ARID2 in hepatocellular carcinoma.

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