Erythroid Differentiation of Mouse Erythroleukemia Cells Results in Reorganization of Protein-DNA Complexes in the Mouse β<sup>Maj</sup> Globin Promoter but not its Distal Enhancer

Reddy, P M; Shen, Che Kun James

doi:10.1128/mcb.13.2.1093-1103.1993

Cited by 6 publications

(1 citation statement)

References 67 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…To specifically knockdown miR-125b-5p in erythroblasts, BALB/c mice were transduced with adeno-associated virus (AAV) serotype 6 which carried beta globin enhancer (GenBank: S73747.1) and promoter (GenBank: GU057255.1) [ 47 ] driving enhanced green fluorescent protein (eGFP) together with either miR-125b sponge (8-repeat antisense-miR-125b-5p) or control (8-repeat scramble nucleotides). The designed enhancer-promoter-eGFP with 8 tandem repeat miR-125b-5p sponges (5′-TCACAAGTTAGGGTCTCAGGGA-3′) or a control (5′-AAGTTTTCAGAAAGCTAACA-3′) [ 48 ] was cloned into AAV by Genechem Co. Ltd. (Shanghai, China).…”

Section: Methodsmentioning

confidence: 99%

The accumulation of miR-125b-5p is indispensable for efficient erythroblast enucleation

Fang

Liang

et al. 2022

Cell Death Dis

View full text Add to dashboard Cite

Erythroblast enucleation is a precisely regulated but not clearly understood process. Polycythemia shows pathological erythroblast enucleation, and we discovered a low miR-125b-5p level in terminal erythroblasts of patients with polycythemia vera (PV) compared to those of healthy controls. Exogenous upregulation of miR-125b-5p levels restored the enucleation rate to normal levels. Direct downregulation of miR-125b-5p in mouse erythroblasts simulated the enucleation issue found in patients with PV, and miR-125b-5p accumulation was found in enucleating erythroblasts, collectively suggesting the importance of miR-125b-5p accumulation for erythroblast enucleation. To elucidate the role of miR-125b-5p in enucleation, gain- and loss-of-function studies were performed. Overexpression of miR-125b-5p improved the enucleation of erythroleukemia cells and primary erythroblasts. Infused erythroblasts with higher levels of miR-125b-5p also exhibited accelerated enucleation. In contrast, miR-125b-5p inhibitors significantly suppressed erythrocyte enucleation. Intracellular imaging revealed that in addition to cytoskeletal assembly and nuclear condensation, miR-125b-5p overexpression resulted in mitochondrial reduction and depolarization. Real-time PCR, western blot analysis, luciferase reporter assays, small molecule inhibitor supplementation and gene rescue assays revealed that Bcl-2, as a direct target of miR-125b-5p, was one of the key mediators of miR-125b-5p during enucleation. Following suppression of Bcl-2, the activation of caspase-3 and subsequent activation of ROCK-1 resulted in cytoskeletal rearrangement and enucleation. In conclusion, this study is the first to reveal the pivotal role of miR-125b-5p in erythroblast enucleation.

show abstract

Section: Methodsmentioning

confidence: 99%

The accumulation of miR-125b-5p is indispensable for efficient erythroblast enucleation

Fang

Liang

et al. 2022

Cell Death Dis

View full text Add to dashboard Cite

show abstract

Activation of β-major globin gene transcription is associated with recruitment of NF-E2 to the β-globin LCR and gene promoter

Sawado

Igarashi

Groudine

2001

Proc. Natl. Acad. Sci. U.S.A.

121

158

View full text Add to dashboard Cite

The mouse ␤-globin gene locus control region (LCR), located upstream of the ␤-globin gene cluster, is essential for the activated transcription of genes in the cluster. The LCR contains multiple binding sites for transactivators, including Maf-recognition elements (MAREs). However, little is known about the specific proteins that bind to these sites or the time at which they bind during erythroid differentiation. We have performed chromatin immunoprecipitation experiments to determine the recruitment of the erythroid-specific transactivator p45 NF-E2͞MafK (p18 NF-E2) heterodimer and small Maf proteins to various regions in the globin gene locus before and after the induction of murine erythroleukemia (MEL) cell differentiation. We report that, before induction, the LCR is occupied by small Maf proteins, and, on erythroid maturation, the NF-E2 complex is recruited to the LCR and the active globin promoters, even though the promoters do not contain MAREs. This differentiation-coupled recruitment of NF-E2 complex correlates with a greater than 100-fold increase in ␤-major globin transcription, but is not associated with a significant change in locus-wide histone H3 acetylation. These findings suggest that the ␤-globin gene locus exists in a constitutively open chromatin conformation before terminal differentiation, and we speculate that recruitment of NF-E2 complex to the LCR and active promoters may be a rate-limiting step in the activation of ␤-globin gene expression.

show abstract

Alterations in Expression and Chromatin Configuration of the Alpha Hemoglobin-Stabilizing Protein Gene in Erythroid Krüppel-Like Factor-Deficient Mice

Pilon

Nilson

Zhou

et al. 2006

Molecular and Cellular Biology

View full text Add to dashboard Cite

Erythroid Krüppel-like factor (EKLF) is an erythroid zinc finger protein identified by its interaction with a CACCC sequence in the ␤-globin promoter, where it establishes local chromatin structure permitting ␤-globin gene transcription. We sought to identify other EKLF target genes and determine the chromatin status of these genes in the presence and absence of EKLF. We identified alpha hemoglobin-stabilizing protein (AHSP) by subtractive hybridization and demonstrated a 95 to 99.9% reduction in AHSP mRNA and the absence of AHSP in EKLF-deficient cells. Chromatin at the AHSP promoter from EKLF-deficient cells lacked a DNase I hypersensitive site and exhibited histone hypoacetylation across the locus compared to hyperacetylation of wild-type chromatin. Wild-type chromatin demonstrated a peak of EKLF binding over a promoter region CACCC box that differs from the EKLF consensus by a nucleotide. In mobility shift assays, the AHSP promoter CACCC site bound EKLF in a manner comparable to the ␤-globin promoter CACCC site, indicating a broader recognition sequence for the EKLF consensus binding site. The AHSP promoter was transactivated by EKLF in K562 cells, which lack EKLF. These results support the hypothesis that EKLF acts as a transcription factor and a chromatin modulator for the AHSP and ␤-globin genes and indicate that EKLF may play similar roles for other erythroid genes.

show abstract

Erythroid Differentiation of Mouse Erythroleukemia Cells Results in Reorganization of Protein-DNA Complexes in the Mouse β^Maj Globin Promoter but not its Distal Enhancer

Cited by 6 publications

References 67 publications

The accumulation of miR-125b-5p is indispensable for efficient erythroblast enucleation

The accumulation of miR-125b-5p is indispensable for efficient erythroblast enucleation

Activation of β-major globin gene transcription is associated with recruitment of NF-E2 to the β-globin LCR and gene promoter

Alterations in Expression and Chromatin Configuration of the Alpha Hemoglobin-Stabilizing Protein Gene in Erythroid Krüppel-Like Factor-Deficient Mice

Contact Info

Product

Resources

About

Erythroid Differentiation of Mouse Erythroleukemia Cells Results in Reorganization of Protein-DNA Complexes in the Mouse βMaj Globin Promoter but not its Distal Enhancer

Cited by 6 publications

References 67 publications

The accumulation of miR-125b-5p is indispensable for efficient erythroblast enucleation

The accumulation of miR-125b-5p is indispensable for efficient erythroblast enucleation

Activation of β-major globin gene transcription is associated with recruitment of NF-E2 to the β-globin LCR and gene promoter

Alterations in Expression and Chromatin Configuration of the Alpha Hemoglobin-Stabilizing Protein Gene in Erythroid Krüppel-Like Factor-Deficient Mice

Contact Info

Product

Resources

About

Erythroid Differentiation of Mouse Erythroleukemia Cells Results in Reorganization of Protein-DNA Complexes in the Mouse β^Maj Globin Promoter but not its Distal Enhancer