scholarly journals The leukemic oncogene EVI1 hijacks a MYC super-enhancer by CTCF-facilitated loops

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Sophie Ottema ◽  
Roger Mulet-Lazaro ◽  
Claudia Erpelinck-Verschueren ◽  
Stanley van Herk ◽  
Marije Havermans ◽  
...  
Keyword(s):  
Chromosomal Rearrangements ◽  
Ctcf Binding ◽  
Ctcf Binding Site ◽  
Common Mechanism ◽  
Docking Site ◽  
Super Enhancer ◽  
Vitro Model ◽  
Ctcf Site ◽  
Promoter Interaction

AbstractChromosomal rearrangements are a frequent cause of oncogene deregulation in human malignancies. Overexpression of EVI1 is found in a subgroup of acute myeloid leukemia (AML) with 3q26 chromosomal rearrangements, which is often therapy resistant. In AMLs harboring a t(3;8)(q26;q24), we observed the translocation of a MYC super-enhancer (MYC SE) to the EVI1 locus. We generated an in vitro model mimicking a patient-based t(3;8)(q26;q24) using CRISPR-Cas9 technology and demonstrated hyperactivation of EVI1 by the hijacked MYC SE. This MYC SE contains multiple enhancer modules, of which only one recruits transcription factors active in early hematopoiesis. This enhancer module is critical for EVI1 overexpression as well as enhancer-promoter interaction. Multiple CTCF binding regions in the MYC SE facilitate this enhancer-promoter interaction, which also involves a CTCF binding site upstream of the EVI1 promoter. We hypothesize that this CTCF site acts as an enhancer-docking site in t(3;8) AML. Genomic analyses of other 3q26-rearranged AML patient cells point to a common mechanism by which EVI1 uses this docking site to hijack enhancers active in early hematopoiesis.

scholarly journals 3'HS1 CTCF binding site in human β-globin locus regulates fetal hemoglobin expression

2021 ◽  
Author(s):  
Pamela Himadewi ◽  
Xue Qing David Wang ◽  
Fan Feng ◽  
Haley Gore ◽  
Yushuai Liu ◽  
...  
Keyword(s):  
Binding Site ◽  
Progenitor Cells ◽  
Globin Gene ◽  
Fetal Hemoglobin ◽  
Ctcf Binding ◽  
Ctcf Binding Site ◽  
Distal Enhancer ◽  
Erythroid Progenitor ◽  
Hematopoietic Stem ◽  
Ctcf Site

Mutations in the adult β-globin gene can lead to a variety of hemoglobinopathies, including sickle cell disease and β-thalassemia. An increase in fetal hemoglobin expression throughout adulthood, a condition named Hereditary Persistence of Fetal Hemoglobin (HPFH), has been found to ameliorate hemoglobinopathies. Deletional HPFH occurs through the excision of a significant portion of the 3 prime end of the β-globin locus, including a CTCF binding site termed 3'HS1. Here, we show that the deletion of this CTCF site alone induces fetal hemoglobin expression in both adult CD34+ hematopoietic stem and progenitor cells and HUDEP-2 erythroid progenitor cells. This induction is driven by the ectopic access of a previously postulated distal enhancer located in the OR52A1 gene downstream of the locus, which can also be insulated by the inversion of the 3'HS1 CTCF site. This suggests that genetic editing of this binding site can have therapeutic implications to treat hemoglobinopathies.

210 DNA METHYLATION AND HYDROXYMETHYLATION ANALYSIS IN A MODEL OF OOCYTE DIFFERENTIAL DEVELOPMENTAL COMPETENCE IN SHEEP

2015 ◽  
Vol 27 (1) ◽  
pp. 195
Author(s):  
L. Masala ◽  
D. Bebbere ◽  
G. P. Burrai ◽  
F. Ariu ◽  
L. Bogliolo ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Embryo Development ◽  
Oocyte Quality ◽  
Developmental Competence ◽  
Ctcf Binding ◽  
Ctcf Binding Site ◽  
Global Methylation ◽  
Tet Genes ◽  
Lower Expression

DNA methylation is an important epigenetic mark that plays a role in gene regulation by the addition of a methyl group to CpG islands in the DNA. Despite being relatively stable in somatic cells, DNA methylation is subject to reprogramming during embryo development and gametogenesis. The aim of this work was to evaluate different aspects of DNA methylation in relation to oocyte quality in the ovine species. A model of differential developmental competence consisting in ovine oocytes and in vitro produced (IVP) blastocysts derived from adult (AD) and prepubertal (PR) donors, was used. The methylation was analysed in terms of: expression of a panel of genes involved in DNA methylation [DNA methyltransferases (DNMTs)] and demethylation [ten-eleven translocation dioxygenases (TET)] in oocytes and blastocysts; global methylation and hydroxymethylation by direct immunofluorescence; locus-specific methylation of 2 imprinted genes by pyrosequencing. Gene relative quantification was performed by RNA reverse transcription followed by real-time PCR. Pools of 10 immature (GV) and in vitro-matured (MII) oocytes and (IVP) blastocysts derived from AD and PR donors (4 replicates per class) were analysed. Lower expression of TET1, TET2, and TET3 was observed in PR GV oocytes (ANOVA; P < 0.05), while no significant differences were found for the enzymes involved in methylation (DNMT1, DNMT3A, DNMT3B; ANOVA; P > 0.05). The levels of all the genes studied showed no significant differences in embryos at blastocyst stage (ANOVA; P > 0.05). Methylation and hydroxymethylation immunostaining were performed in GV and MII oocytes using anti-5-methylcytosine mouse mAb and 5-hydroxymethylcytosine rabbit pAB. High levels of DNA methylation were observed in both AD and PR GV and MII oocytes, while hydroxymethylation immunopositivity was scattered evident throughout the gamete chromatin. Pyrosequencing of bisulfite converted DNA was used to determine the methylation status within differentially methylated regions (DMR) of maternally imprinted H19 (CTCF binding site IV; 11 CpG sites) and paternally imprinted IGF2R (17CpG sites within intron 2). No differences were observed between classes of oocytes for each gene (pools of 40 oocytes per replicate, 3 replicates per class; ANOVA; P > 0.05). Our work shows no differences in the expression of the enzymes involved in methylation, in accordance with the results of global and locus specific methylation analysis. Conversely, we observed lower expression of the TET genes in PR GV oocytes (ANOVA; P > 0.05). TET1, TET2, and TET3, whose expression has never been studied in ovine, generate 5-hydroxymethlcytosine (5hmC) by oxidation of 5-methylcytosine (5mC), and are involved in active DNA demethylation during early embryo development. Our observation of lower expression of the TET genes in lower competence PR GV oocytes suggests that epigenetic mechanisms may affect oocyte quality and paves the way to better understand methylation dynamics during sheep pre-implantation development.

scholarly journals Canonical WNT signaling-dependent gating of MYC requires a noncanonical CTCF function at a distal binding site

2022 ◽  
Vol 13 (1) ◽  
Author(s):  
Ilyas Chachoua ◽  
Ilias Tzelepis ◽  
Hao Dai ◽  
Jia Pei Lim ◽  
Anna Lewandowska-Ronnegren ◽  
...  
Keyword(s):  
Wnt Signaling ◽  
Binding Site ◽  
Nuclear Export ◽  
Nuclear Architecture ◽  
Ctcf Binding ◽  
Ctcf Binding Site ◽  
Nuclear Pore ◽  
Super Enhancer ◽  
Hct 116 Cells ◽  
Canonical Wnt

AbstractAbnormal WNT signaling increases MYC expression in colon cancer cells in part via oncogenic super-enhancer-(OSE)-mediated gating of the active MYC to the nuclear pore in a poorly understood process. We show here that the principal tenet of the WNT-regulated MYC gating, facilitating nuclear export of the MYC mRNA, is regulated by a CTCF binding site (CTCFBS) within the OSE to confer growth advantage in HCT-116 cells. To achieve this, the CTCFBS directs the WNT-dependent trafficking of the OSE to the nuclear pore from intra-nucleoplasmic positions in a stepwise manner. Once the OSE reaches a peripheral position, which is triggered by a CTCFBS-mediated CCAT1 eRNA activation, its final stretch (≤0.7 μm) to the nuclear pore requires the recruitment of AHCTF1, a key nucleoporin, to the CTCFBS. Thus, a WNT/ß-catenin-AHCTF1-CTCF-eRNA circuit enables the OSE to promote pathological cell growth by coordinating the trafficking of the active MYC gene within the 3D nuclear architecture.

scholarly journals Alteration of CTCF-associated chromatin neighborhood inhibits TAL1-driven oncogenic transcription program and leukemogenesis

2020 ◽  
Vol 48 (6) ◽  
pp. 3119-3133 ◽  
Author(s):  
Ying Li ◽  
Ziwei Liao ◽  
Huacheng Luo ◽  
Aissa Benyoucef ◽  
Yuanyuan Kang ◽  
...  
Keyword(s):  
Chromatin Accessibility ◽  
Jurkat Cells ◽  
Ctcf Binding ◽  
Ctcf Binding Site ◽  
Dependent Manner ◽  
Range Interaction ◽  
Chromatin Boundary ◽  
Transcription Networks ◽  
Transcription Program ◽  
Promoter Interaction

Abstract Aberrant activation of the TAL1 is associated with up to 60% of T-ALL cases and is involved in CTCF-mediated genome organization within the TAL1 locus, suggesting that CTCF boundary plays a pathogenic role in T-ALL. Here, we show that −31-Kb CTCF binding site (−31CBS) serves as chromatin boundary that defines topologically associating domain (TAD) and enhancer/promoter interaction required for TAL1 activation. Deleted or inverted −31CBS impairs TAL1 expression in a context-dependent manner. Deletion of −31CBS reduces chromatin accessibility and blocks long-range interaction between the +51 erythroid enhancer and TAL1 promoter-1 leading to inhibition of TAL1 expression in erythroid cells, but not T-ALL cells. However, in TAL1-expressing T-ALL cells, the leukemia-prone TAL1 promoter-IV specifically interacts with the +19 stem cell enhancer located 19 Kb downstream of TAL1 and this interaction is disrupted by the −31CBS inversion in T-ALL cells. Inversion of −31CBS in Jurkat cells alters chromatin accessibility, histone modifications and CTCF-mediated TAD leading to inhibition of TAL1 expression and TAL1-driven leukemogenesis. Thus, our data reveal that −31CBS acts as critical regulator to define +19-enhancer and the leukemic prone promoter IV interaction for TAL1 activation in T-ALL. Manipulation of CTCF boundary can alter TAL1 TAD and oncogenic transcription networks in leukemogenesis.

scholarly journals A Variably Occupied CTCF Binding Site in the Ultrabithorax Gene in the Drosophila Bithorax Complex

2014 ◽  
Vol 35 (1) ◽  
pp. 318-330 ◽  
Author(s):  
Jose Paolo Magbanua ◽  
Estelle Runneburger ◽  
Steven Russell ◽  
Robert White
Keyword(s):  
Imaginal Disc ◽  
Normal Function ◽  
Ctcf Binding ◽  
Ctcf Binding Site ◽  
Content Type ◽  
Variable Site ◽  
Binding Factor ◽  
Context Specific ◽  
Transposon Insertions ◽  
Ctcf Site

Although the majority of genomic binding sites for the insulator protein CCCTC-binding factor (CTCF) are constitutively occupied, a subset show variable occupancy. Such variable sites provide an opportunity to assess context-specific CTCF functions in gene regulation. Here, we have identified a variably occupied CTCF site in theDrosophila Ultrabithorax(Ubx) gene. This site is occupied in tissues whereUbxis active (third thoracic leg imaginal disc) but is not bound in tissues where theUbxgene is repressed (first thoracic leg imaginal disc). Using chromatin conformation capture, we show that this site preferentially interacts with theUbxpromoter region in the active state. The site lies close toUbxenhancer elements and is also close to the locations of severalgypsytransposon insertions that disruptUbxexpression, leading to thebxmutant phenotype.gypsyinsertions carry the Su(Hw)-dependentgypsyinsulator and were found to affect both CTCF binding at the variable site and the chromatin topology. This suggests that insertion of thegypsyinsulator in this region interferes with CTCF function and supports a model for the normal function of the variable CTCF site as a chromatin loop facilitator, promoting interaction betweenUbxenhancers and theUbxtranscription start site.

scholarly journals Analysis of Sequence Upstream of the Endogenous H19 Gene Reveals Elements Both Essential and Dispensable for Imprinting

2002 ◽  
Vol 22 (8) ◽  
pp. 2450-2462 ◽  
Author(s):  
Joanne L. Thorvaldsen ◽  
Mellissa R. W. Mann ◽  
Okechukwu Nwoko ◽  
Kristen L. Duran ◽  
Marisa S. Bartolomei
Keyword(s):  
Repetitive Elements ◽  
Ctcf Binding ◽  
Ctcf Binding Site ◽  
Insulator Activity ◽  
Binding Factor ◽  
Additional Sequence ◽  
H19 Gene ◽  
Full Activation ◽  
Ctcf Site

ABSTRACT Imprinting of the linked and oppositely expressed mouse H19 and Igf2 genes requires a 2-kb differentially methylated domain (DMD) that is located 2 kb upstream of H19. This element is postulated to function as a methylation-sensitive insulator. Here we test whether an additional sequence 5′ of H19 is required for H19 and Igf2 imprinting. Because repetitive elements have been suggested to be important for genomic imprinting, the requirement of a G-rich repetitive element that is located immediately 3′ to the DMD was first tested in two targeted deletions: a 2.9-kb deletion (ΔDMDΔG) that removes the DMD and G-rich repeat and a 1.3-kb deletion (ΔG) removing only the latter. There are also four 21-bp GC-rich repetitive elements within the DMD that bind the insulator-associated CTCF (CCCTC-binding factor) protein and are implicated in mediating methylation-sensitive insulator activity. As three of the four repeats of the 2-kb DMD were deleted in the initial 1.6-kb ΔDMD allele, we analyzed a 3.8-kb targeted allele (Δ3.8kb-5′H19), which deletes the entire DMD, to test the function of the fourth repeat. Comparative analysis of the 5′ deletion alleles reveals that (i) the G-rich repeat element is dispensable for imprinting, (ii) the ΔDMD and ΔDMDΔG alleles exhibit slightly more methylation upon paternal transmission, (iii) removal of the 5′ CTCF site does not further perturb H19 and Igf2 imprinting, suggesting that one CTCF-binding site is insufficient to generate insulator activity in vivo, (iv) the DMD sequence is required for full activation of H19 and Igf2, and (v) deletion of the DMD disrupts H19 and Igf2 expression in a tissue-specific manner.

scholarly journals HTLV-1 CTCF-binding site is dispensable for in vitro immortalization and persistent infection in vivo

Retrovirology ◽  
2019 ◽  
Vol 16 (1) ◽  
Author(s):  
Michael P. Martinez ◽  
Xiaogang Cheng ◽  
Ancy Joseph ◽  
Jacob Al-Saleem ◽  
Amanda R. Panfil ◽  
...  
Keyword(s):  
Gene Expression ◽  
T Cell ◽  
Antibody Response ◽  
Specific Antibody ◽  
Ctcf Binding ◽  
Ctcf Binding Site ◽  
Cell Leukemia ◽  
Specific Antibody Response

Abstract Background Human T-cell leukemia virus type 1 (HTLV-1) is the etiologic agent of adult T-cell leukemia/lymphoma (ATL) and the neurological disorder HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). The exact mechanism(s) through which latency and disease progression are regulated are not fully understood. CCCTC-binding factor (CTCF) is an 11-zinc finger, sequence-specific, DNA-binding protein with thousands of binding sites throughout mammalian genomes. CTCF has been shown to play a role in organization of higher-order chromatin structure, gene expression, genomic imprinting, and serve as a barrier to epigenetic modification. A viral CTCF-binding site (vCTCF-BS) was previously identified within the overlapping p12 (sense) and Hbz (antisense) genes of the HTLV-1 genome. Thus, upon integration, HTLV-1 randomly inserts a vCTCF-BS into the host genome. vCTCF-BS studies to date have focused primarily on HTLV-1 chronically infected or tumor-derived cell lines. In these studies, HTLV-1 was shown to alter the structure and transcription of the surrounding host chromatin through the newly inserted vCTCF-BS. However, the effects of CTCF binding in the early stages of HTLV-1 infection remains unexplored. This study examines the effects of the vCTCF-BS on HTLV-1-induced in vitro immortalization and in vivo viral persistence in infected rabbits. Results HTLV-1 and HTLV-1∆CTCF LTR-transactivation, viral particle production, and immortalization capacity were comparable in vitro. The total lymphocyte count, proviral load, and Hbz gene expression were not significantly different between HTLV-1 and HTLV-1∆CTCF-infected rabbits throughout a 12 week study. However, HTLV-1∆CTCF-infected rabbits displayed a significantly decreased HTLV-1-specific antibody response compared to HTLV-1-infected rabbits. Conclusions Mutation of the HTLV-1 vCTCF-BS does not significantly alter T-lymphocyte transformation capacity or early in vivo virus persistence, but results in a decreased HTLV-1-specific antibody response during early infection in rabbits. Ultimately, understanding epigenetic regulation of HTLV-1 gene expression and pathogenesis could provide meaningful insights into mechanisms of immune evasion and novel therapeutic targets.

scholarly journals The full oncogenic activity of Ret/ptc2 depends on tyrosine 539, a docking site for phospholipase Cgamma.

1996 ◽  
Vol 16 (5) ◽  
pp. 2151-2163 ◽  
Author(s):  
M G Borrello ◽  
L Alberti ◽  
E Arighi ◽  
I Bongarzone ◽  
C Battistini ◽  
...  
Keyword(s):  
Chromosomal Rearrangements ◽  
Docking Site ◽  
Kinetic Measurements ◽  
Downstream Effector ◽  
Gene Coding ◽  
Oncogenic Activity ◽  
Affinity Interaction ◽  
Nih 3T3

RET/PTC oncogenes, generated by chromosomal rearrangements in papillary thyroid carcinomas, are constitutively activated versions of proto-RET, a gene coding for a receptor-type tyrosine kinase (TK) whose ligand is still unknown. RET/PTCs encode fusion proteins in which proto-RET TK and C-terminal domains are fused to different donor genes. The respective Ret/ptc oncoproteins display constitutive TK activity and tyrosine phosphorylation. We found that Ret/ptcs associate with and phosphorylate the SH2-containing transducer phospholipase Cgamma (PLCgamma). Two putative PLCgamma docking sites, Tyr-505 and Tyr-539, have been identified on Ret/ptc2 by competition experiments using phosphorylated peptides modelled on Ret sequence. Transfection experiments and biochemical analysis using Tyr-->Phe mutants of Ret/ptc2 allowed us to rule out Tyr-505 and to identify Tyr-539 as a functional PLCgamma docking site in vivo. Moreover, kinetic measurements showed that Tyr-539 is able to mediate high-affinity interaction with PLCgamma. Mutation of Tyr-539 resulted in a drastically reduced oncogenic activity of Ret/ptc2 on NIH 3T3 cells (75 to 90% reduction) both in vitro and in vivo, which correlates with impaired ability of Ret/ptc2 to activate PLCgamma. In conclusion, this paper demonstrates that Tyr-539 of Ret/ptc2 (Tyr-761 on the proto-RET product) is an essential docking site for the full transforming potential of the oncogene. In addition, the present data identify PLCgamma as a downstream effector of Ret/ptcs and suggest that this transducing molecule could play a crucial role in neoplastic signalling triggered by Ret/ptc oncoproteins.

scholarly journals 3'HS1 CTCF binding site in human β-globin locus regulates fetal hemoglobin expression

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Pamela Himadewi ◽  
Xue Qing David Wang ◽  
Fan Feng ◽  
Haley Gore ◽  
Yushuai Liu ◽  
...  
Keyword(s):  
Binding Site ◽  
Progenitor Cells ◽  
Globin Gene ◽  
Fetal Hemoglobin ◽  
Ctcf Binding ◽  
Ctcf Binding Site ◽  
Distal Enhancer ◽  
Erythroid Progenitor ◽  
Hematopoietic Stem ◽  
Ctcf Site

Mutations in the adult β-globin gene can lead to a variety of hemoglobinopathies, including sickle cell disease and β-thalassemia. An increase in fetal hemoglobin expression throughout adulthood, a condition named Hereditary Persistence of Fetal Hemoglobin (HPFH), has been found to ameliorate hemoglobinopathies. Deletional HPFH occurs through the excision of a significant portion of the 3' end of the β-globin locus, including a CTCF binding site termed 3'HS1. Here, we show that the deletion of this CTCF site alone induces fetal hemoglobin expression in both adult CD34+ hematopoietic stem and progenitor cells and HUDEP-2 erythroid progenitor cells. This induction is driven by the ectopic access of a previously postulated distal enhancer located in the OR52A1 gene downstream of the locus, which can also be insulated by the inversion of the 3'HS1 CTCF site. This suggests that genetic editing of this binding site can have therapeutic implications to treat hemoglobinopathies.

An in vitro model for investigation of vitamin A effects on wound healing

2021 ◽  
pp. 1-6
Author(s):  
Hoda Keshmiri Neghab ◽  
Mohammad Hasan Soheilifar ◽  
Gholamreza Esmaeeli Djavid
Keyword(s):  
Wound Healing ◽  
Endothelial Cell ◽  
Vitamin A ◽  
In Vitro Model ◽  
Cellular Proliferation ◽  
Endothelial Cell Migration ◽  
Normal Fibroblast ◽  
Anti Inflammatory ◽  
Vitro Model

Abstract. Wound healing consists of a series of highly orderly overlapping processes characterized by hemostasis, inflammation, proliferation, and remodeling. Prolongation or interruption in each phase can lead to delayed wound healing or a non-healing chronic wound. Vitamin A is a crucial nutrient that is most beneficial for the health of the skin. The present study was undertaken to determine the effect of vitamin A on regeneration, angiogenesis, and inflammation characteristics in an in vitro model system during wound healing. For this purpose, mouse skin normal fibroblast (L929), human umbilical vein endothelial cell (HUVEC), and monocyte/macrophage-like cell line (RAW 264.7) were considered to evaluate proliferation, angiogenesis, and anti-inflammatory responses, respectively. Vitamin A (0.1–5 μM) increased cellular proliferation of L929 and HUVEC (p < 0.05). Similarly, it stimulated angiogenesis by promoting endothelial cell migration up to approximately 4 fold and interestingly tube formation up to 8.5 fold (p < 0.01). Furthermore, vitamin A treatment was shown to decrease the level of nitric oxide production in a dose-dependent effect (p < 0.05), exhibiting the anti-inflammatory property of vitamin A in accelerating wound healing. These results may reveal the therapeutic potential of vitamin A in diabetic wound healing by stimulating regeneration, angiogenesis, and anti-inflammation responses.

Sign in / Sign up

Export Citation Format

Share Document