Wild-type egr1/Krox24 promotes and dominant-negative mutants inhibit, pluripotent differentiation of p19 embryonal carcinoma cells

Lanoix, Joël; Mullick, Alaka; He, Yulan; Bravo, Rodrigo; Skup, Daniel

doi:10.1038/sj.onc.1202166

Cited by 13 publications

(16 citation statements)

References 42 publications

(64 reference statements)

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“…The loss of Egr1 has effects in both hematopoietic and neural lineages and appears to be a potent inducer of differentiation in several multipotent cell types. In P19 cells, tight control of Egr1 is essential for maintaining the pluripotent state (22,24).…”

Section: Discussionmentioning

confidence: 99%

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The Histone Demethylase KDM5b/JARID1b Plays a Role in Cell Fate Decisions by Blocking Terminal Differentiation

Dey

Stalker

Schnerch

et al. 2008

Molecular and Cellular Biology

116

111

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The histone demethylase lysine demethylase 5b (KDM5b) specifically demethylates lysine 4 of histone H3 (meH3K4), thereby repressing gene transcription. KDM5b regulates cell cycle control genes in cancer and is expressed in the early epiblast. This suggests that KDM5b plays a developmental role by maintaining uncommitted progenitors. Here we show that transient overexpression of KDM5b in embryonic stem cells decreases the expression of at least three different modulators of cell fate decisions, Egr1, p27 KIP1 , and BMI1, by demethylation of their promoters. Constitutively increased KDM5b expression results in an increased mitotic rate and a decreased global 3meH3K4 but no change in cell identity. Results of two separate differentiation assays, neural differentiation and embryoid body EB (EB) formation, showed that KDM5b reduced the terminally differentiated cells and increased proliferating progenitors. These were achieved by two mechanisms, blocking of the upregulation of cell lineage markers and maintenance of cyclins, that allowed cells to escape differentiation and remain uncommitted. Additionally, EBs maintain high levels of Oct4 and Nanog and can be dissociated to reestablish highly proliferative cultures. The persistence of uncommitted progenitors may be due to the direct regulation of the Tcf/Lef family member mTcf3/hTcf7L1, an upstream regulator of Nanog expression. These findings demonstrate a role for KDM5b in the choice between proliferation and differentiation during development.Transcriptional control is a dynamic process, during which several different histone residues are modified to change RNA polymerase's ability to access the transcriptional start site (19,42). A key component in this process is the methylation of histone H3 lysine 4 (H3K4). Methylation of H3K4 is a key regulator of RNA polymerase binding to active genes (41) and of transcription factor binding within promoter elements (43). The ability of this epigenetic mark to control multiple points in transcription suggests that modulation of H3K4 methylation plays a role in both the activation and the repression of genes.A key aspect of H3K4 methylation is how this epigenetic mark is removed, thereby reducing RNA polymerase's localization to the specific genes. This loss of methyl H3K4

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Section: Discussionmentioning

confidence: 99%

“…In P19 EC cells, increased Egr1 levels cause spontaneous differentiation, whereas dominant-negative Egr1 strongly blocks differentiation (24). This suggests a conserved role for Egr1 in differentiation.…”

Section: And Mescmentioning

confidence: 92%

The Histone Demethylase KDM5b/JARID1b Plays a Role in Cell Fate Decisions by Blocking Terminal Differentiation

Dey

Stalker

Schnerch

et al. 2008

Molecular and Cellular Biology

116

111

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show abstract

“…Egr proteins can repress transcription by displacement of Sp1, a well characterized, ubiquitous zinc fingercontaining, DNA-binding protein (47). Importantly, this repression-by-displacement mechanism appears to require more than just the DNA-binding domain of the Egr protein, because it can be blocked by overexpression of an Egr1 construct lacking the Nterminal regulatory domain (48). Genes whose expression is downregulated in this manner are distinguished by the absence of a proximal TATA or CAAT box and the presence of overlapping Sp1/Egr1 binding sites (18,47).…”

Section: Discussionmentioning

confidence: 99%

Early Growth Response Transcription Factors Are Required for Development of CD4−CD8− Thymocytes to the CD4+CD8+ Stage

Carleton

Haks

Smeele

et al. 2002

The Journal of Immunology

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Progression of immature CD4−CD8− thymocytes beyond the β-selection checkpoint to the CD4+CD8+ stage requires activation of the pre-TCR complex; however, few of the DNA-binding proteins that serve as molecular effectors of those pre-TCR signals have been identified. We demonstrate in this study that members of the early growth response (Egr) family of transcription factors are critical effectors of the signals that promote this developmental transition. Specifically, the induction of three Egr family members (Egr1, 2, and 3) correlates with pre-TCR activation and development of CD4−CD8− thymocytes beyond the β-selection checkpoint. Enforced expression of each of these Egr factors is able to bypass the block in thymocyte development associated with defective pre-TCR function. However, Egr family members may play somewhat distinct roles in promoting thymocyte development, because there are differences in the genes modulated by enforced expression of particular Egr factors. Finally, interfering with Egr function using dominant-negative proteins disrupts thymocyte development from the CD4−CD8− to the CD4+CD8+ stage. Taken together, these data demonstrate that the Egr proteins play an essential role in executing the differentiation program initiated by pre-TCR signaling.

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“…Constructs, Transfections, and Reporter Assays-The PGK-puro vector, WT Egr-1/Krox-24, and dominant-negative mutants (WT minus the zinc finger region and the zinc finger region only) were a kind gift from Dr. D. Skup, and the construction of the plasmids has been described previously (28). Briefly, human egr-1/krox-24 cDNA was removed from the pMexNeoKrox-24 vector and inserted into the EcoRI site of the PGK-puro vector (puromycin for selection), which utilizes the PGK promoter to drive transcription, and thus generated the pPGK-Egr-1/ Krox-24 expression plasmid.…”

Section: Methodsmentioning

confidence: 99%

“…The oligonucleotides were used to amplify the zinc finger region from pMexNeoKrox-24, and the resulting fragment was then cloned into BamHI/ XbaI-digested PGK vector. Sequencing confirmed that the insert was correct and in-frame (28). Promoter constructs containing 1395 bp of the egr-1/krox-24 promoter region along with Ϫ925, Ϫ636, Ϫ459, Ϫ252, and Ϫ77 deletion mutants were kindly provided by Dr. D. Skup, whereas the pTNFϪ615CAT construct was a gift from Dr.…”

Section: Methodsmentioning

confidence: 99%

Early Growth Response Factor-1 Mediates Prostaglandin E2-dependent Transcriptional Suppression of Cytokine-induced Tumor Necrosis Factor-α Gene Expression in Human Macrophages and Rheumatoid Arthritis-affected Synovial Fibroblasts

Faour

Alaaeddine

Mancini

et al. 2005

Journal of Biological Chemistry

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Tumor necrosis factor-␣ (TNF-␣) is a pleiotropic proinflammatory cytokine that modulates a broad range of inflammatory and immunological processes. We have investigated the potential immunomodulatory properties of prostaglandin E 2 (PGE 2 ) by examining the molecular mechanism by which the eicosanoid suppresses T-cell-derived interleukin-17 (IL-17)-induced TNF-␣ mRNA expression and protein synthesis in human macrophages and rheumatoid arthritis-affected synovial fibroblasts. Initial studies confirmed that PGE 2 induces egr-1 mRNA expression and protein synthesis by restricted SAPK2/p38 MAPKdependent activating transcription factor-2 (ATF-2) dimer transactivation of the egr-1 promoter as judged by studies using wild-type (WT) and deletion mutant egr-1 promoter constructs, Northern and Western blotting, and standard and supershift electrophoretic mobility shift analyses. Using human leukemic monocytic THP-1 cells stably transfected with WT and dominant-negative mutant expression constructs of Egr-1, cotransfected or not with a WT pTNF؊615SVOCAT construct, we observed that PGE 2 inhibition of IL-17-stimulated TNF-␣ mRNA expression and promoter activity was dependent on Egr-1 expression, as mutants of Egr-1, alone or in combination, markedly abrogated any inhibitory effect of PGE 2 . Standard and supershift electrophoretic mobility shift analysis, signaling "decoy" overexpression studies, and pTNF؊615SVOCAT promoter assays using WT and mutant promoter constructs revealed that IL-17-up-regulated promoter activity was largely dependent on ATF-2/c-Jun transactivation. PGE 2 suppression of IL-17-induced ATF-2/c-Jun transactivation and DNA binding was dependent on Egr-1-mediated inhibition of induced c-Jun expression. We suggest that egr-1 is an immediate-early PGE 2 target gene that may be a key regulatory factor in mediating eicosanoid control of genes involved in the immune and inflammatory responses.

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Wild-type egr1/Krox24 promotes and dominant-negative mutants inhibit, pluripotent differentiation of p19 embryonal carcinoma cells

Cited by 13 publications

References 42 publications

The Histone Demethylase KDM5b/JARID1b Plays a Role in Cell Fate Decisions by Blocking Terminal Differentiation

The Histone Demethylase KDM5b/JARID1b Plays a Role in Cell Fate Decisions by Blocking Terminal Differentiation

Early Growth Response Transcription Factors Are Required for Development of CD4−CD8− Thymocytes to the CD4+CD8+ Stage

Early Growth Response Factor-1 Mediates Prostaglandin E2-dependent Transcriptional Suppression of Cytokine-induced Tumor Necrosis Factor-α Gene Expression in Human Macrophages and Rheumatoid Arthritis-affected Synovial Fibroblasts

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