Journal articles on the topic 'NF-E2'
To see the other types of publications on this topic, follow the link: NF-E2.
Create a spot-on reference in APA, MLA, Chicago, Harvard, and other styles
Consult the top 50 journal articles for your research on the topic 'NF-E2.'
Next to every source in the list of references, there is an 'Add to bibliography' button. Press on it, and we will generate automatically the bibliographic reference to the chosen work in the citation style you need: APA, MLA, Harvard, Chicago, Vancouver, etc.
You can also download the full text of the academic publication as pdf and read online its abstract whenever available in the metadata.
Browse journal articles on a wide variety of disciplines and organise your bibliography correctly.
1
Ney, P. A., N. C. Andrews, S. M. Jane, B. Safer, M. E. Purucker, S. Weremowicz, C. C. Morton, S. C. Goff, S. H. Orkin, and A. W. Nienhuis. "Purification of the human NF-E2 complex: cDNA cloning of the hematopoietic cell-specific subunit and evidence for an associated partner." Molecular and Cellular Biology 13, no. 9 (September 1993): 5604–12. http://dx.doi.org/10.1128/mcb.13.9.5604.
Full textAbstract:
The human globin locus control region-binding protein, NF-E2, was purified by DNA affinity chromatography. Its tissue-specific component, p45 NF-E2, was cloned by use of a low-stringency library screen with murine p45 NF-E2 cDNA (N. C. Andrews, H. Erdjument-Bromage, M. B. Davidson, P. Tempst, and S. H. Orkin, Nature [London] 362:722-728, 1993). The human p45 NF-E2 gene was localized to chromosome 12q13 by fluorescent in situ hybridization. Human p45 NF-E2 and murine p45 NF-E2 are highly homologous basic region-leucine zipper (bZIP) proteins with identical DNA-binding domains. Immunoprecipitation experiments demonstrated that p45 NF-E2 is associated in vivo with an 18-kDa protein (p18). Because bZIP proteins bind DNA as dimers, we infer that native NF-E2 must be a heterodimer of 45- and 18-kDa subunits. Although AP-1 and CREB copurified with NF-E2, no evidence was found for heterodimer formation between p45 NF-E2 and proteins other than p18. Thus, p18 appears to be the sole specific partner of p45 NF-E2 in erythroid cells. Cloning of human p45 NF-E2 should permit studies of the role of NF-E2 in globin gene regulation and erythroid differentiation.
2
Ney, P. A., N. C. Andrews, S. M. Jane, B. Safer, M. E. Purucker, S. Weremowicz, C. C. Morton, S. C. Goff, S. H. Orkin, and A. W. Nienhuis. "Purification of the human NF-E2 complex: cDNA cloning of the hematopoietic cell-specific subunit and evidence for an associated partner." Molecular and Cellular Biology 13, no. 9 (September 1993): 5604–12. http://dx.doi.org/10.1128/mcb.13.9.5604-5612.1993.
Full textAbstract:
The human globin locus control region-binding protein, NF-E2, was purified by DNA affinity chromatography. Its tissue-specific component, p45 NF-E2, was cloned by use of a low-stringency library screen with murine p45 NF-E2 cDNA (N. C. Andrews, H. Erdjument-Bromage, M. B. Davidson, P. Tempst, and S. H. Orkin, Nature [London] 362:722-728, 1993). The human p45 NF-E2 gene was localized to chromosome 12q13 by fluorescent in situ hybridization. Human p45 NF-E2 and murine p45 NF-E2 are highly homologous basic region-leucine zipper (bZIP) proteins with identical DNA-binding domains. Immunoprecipitation experiments demonstrated that p45 NF-E2 is associated in vivo with an 18-kDa protein (p18). Because bZIP proteins bind DNA as dimers, we infer that native NF-E2 must be a heterodimer of 45- and 18-kDa subunits. Although AP-1 and CREB copurified with NF-E2, no evidence was found for heterodimer formation between p45 NF-E2 and proteins other than p18. Thus, p18 appears to be the sole specific partner of p45 NF-E2 in erythroid cells. Cloning of human p45 NF-E2 should permit studies of the role of NF-E2 in globin gene regulation and erythroid differentiation.
3
Wang, Wei, Sven Schwemmers, Elizabeth O. Hexner, and Heike L. Pahl. "AML1 is overexpressed in patients with myeloproliferative neoplasms and mediates JAK2V617F-independent overexpression of NF-E2." Blood 116, no. 2 (July 15, 2010): 254–66. http://dx.doi.org/10.1182/blood-2009-11-254664.
Full textAbstract:
Abstract The transcription factor NF-E2 is overexpressed in the majority of patients with polycythemia vera (PV). Concomitantly, 95% of these patients carry the JAK2V617F mutation. Although NF-E2 levels correlate with JAK2V671F allele burden in some PV cohorts, the molecular mechanism causing aberrant NF-E2 expression has not been described. Here we show that NF-E2 expression is also increased in patients with essential thrombocythemia and primary myelofibrosis independent of the presence of the JAK2V617F mutation. Characterization of the NF-E2 promoter revealed multiple functional binding sites for AML1/RUNX-1. Chromatin immunoprecipitation demonstrated AML1 binding to the NF-E2 promoter in vivo. Moreover, AML1 binding to the NF-E2 promoter was significantly increased in granulocytes from PV patients compared with healthy controls. AML1 mRNA expression was elevated in patients with PV, essential thrombocythemia, and primary myelofibrosis both in the presence and absence of JAK2V617F. In addition, AML1 and NF-E2 expression were highly correlated. RNAi-mediated suppression of either AML1 or of its binding partner CBF-β significantly decreased NF-E2 expression. Moreover, expression of the leukemic fusion protein AML/ETO drastically decreased NF-E2 protein levels. Our data identify NF-E2 as a novel AML1 target gene and delineate a role for aberrant AML1 expression in mediating elevated NF-E2 expression in MPN patients.
4
Martin, Florence, Jan M. van Deursen, Ramesh A. Shivdasani, Carl W. Jackson, Amber G. Troutman, and Paul A. Ney. "Erythroid Maturation and Globin Gene Expression in Mice With Combined Deficiency of NF-E2 and Nrf-2." Blood 91, no. 9 (May 1, 1998): 3459–66. http://dx.doi.org/10.1182/blood.v91.9.3459.
Full textAbstract:
Abstract NF-E2 binding sites, located in distant regulatory sequences, may be important for high level α- and β-globin gene expression. Surprisingly, targeted disruption of each subunit of NF-E2 has either little or no effect on erythroid maturation in mice. For p18 NF-E2, this lack of effect is due, at least in part, to the presence of redundant proteins. For p45 NF-E2, one possibility is that NF-E2–related factors, Nrf-1 or Nrf-2, activate globin gene expression in the absence of NF-E2. To test this hypothesis for Nrf-2, we disrupted the Nrf-2 gene by homologous recombination. Nrf-2–deficient mice had no detectable hematopoietic defect. In addition, no evidence was found for reciprocal upregulation of NF-E2 or Nrf-2 protein in fetal liver cells deficient for either factor. Fetal liver cells deficient for both NF-E2 and Nrf-2 expressed normal levels of α- and β-globin. Mature mice with combined deficiency of NF-E2 and Nrf-2 did not exhibit a defect in erythroid maturation beyond that seen with loss of NF-E2 alone. Thus, the presence of a mild erythroid defect in NF-E2–deficient mice is not the result of compensation by Nrf-2.
5
Martin, Florence, Jan M. van Deursen, Ramesh A. Shivdasani, Carl W. Jackson, Amber G. Troutman, and Paul A. Ney. "Erythroid Maturation and Globin Gene Expression in Mice With Combined Deficiency of NF-E2 and Nrf-2." Blood 91, no. 9 (May 1, 1998): 3459–66. http://dx.doi.org/10.1182/blood.v91.9.3459.3459_3459_3466.
Full textAbstract:
NF-E2 binding sites, located in distant regulatory sequences, may be important for high level α- and β-globin gene expression. Surprisingly, targeted disruption of each subunit of NF-E2 has either little or no effect on erythroid maturation in mice. For p18 NF-E2, this lack of effect is due, at least in part, to the presence of redundant proteins. For p45 NF-E2, one possibility is that NF-E2–related factors, Nrf-1 or Nrf-2, activate globin gene expression in the absence of NF-E2. To test this hypothesis for Nrf-2, we disrupted the Nrf-2 gene by homologous recombination. Nrf-2–deficient mice had no detectable hematopoietic defect. In addition, no evidence was found for reciprocal upregulation of NF-E2 or Nrf-2 protein in fetal liver cells deficient for either factor. Fetal liver cells deficient for both NF-E2 and Nrf-2 expressed normal levels of α- and β-globin. Mature mice with combined deficiency of NF-E2 and Nrf-2 did not exhibit a defect in erythroid maturation beyond that seen with loss of NF-E2 alone. Thus, the presence of a mild erythroid defect in NF-E2–deficient mice is not the result of compensation by Nrf-2.
6
Kotkow, K. J., and S. H. Orkin. "Dependence of globin gene expression in mouse erythroleukemia cells on the NF-E2 heterodimer." Molecular and Cellular Biology 15, no. 8 (August 1995): 4640–47. http://dx.doi.org/10.1128/mcb.15.8.4640.
Full textAbstract:
High-level, tissue-specific expression of the beta-globin genes requires the presence of an upstream locus control region (LCR). The overall enhancer activity of the beta-globin complex LCR (beta-LCR) is dependent on the integrity of the tandem NF-E2 sites of HS-2. The NF-E2 protein which binds these sites is a heterodimeric basic leucine zipper protein composed of a tissue-specific subunit, p45 NF-E2, and a smaller subunit, p18 NF-E2, that is widely expressed. In these studies, we sought to investigate the role of NF-E2 in globin expression. We show that expression of a dominant-negative mutant p18 greatly reduces the amount of functional NF-E2 complex in the cell. Reduced levels of both alpha- and beta-globin were associated with the lower levels of NF-E2 activity in this cell line. Globin expression was fully restored upon the introduction of a tethered p45-p18 heterodimer. We also examined CB3 cells, a mouse erythroleukemia (MEL) cell line that does not express endogenous p45 NF-E2, and demonstrated that the restoration of globin gene expression was dependent upon the levels of expressed tethered NF-E2 heterodimer. Results of DNase I hypersensitivity mapping and in vivo footprinting assays showed no detectable chromatin alterations in beta-LCR HS-2 due to loss of NF-E2. Finally, we examined the specificity of NF-E2 for globin gene expression in MEL cells. These experiments indicate a critical role for the amino-terminal domain of p45 NF-E2 and show that a related protein, LCRF1, is unable to restore globin gene expression in p45 NF-E2-deficient cells. From these results, we conclude that NF-E2 is specifically required for high level goblin gene expression in MEL cells.
7
Jutzi, Jonas S., Ruzhica Bogeska, Gorica Nikoloski, Corina A. Schmid, Thalia S. Seeger, Frank Stegelmann, Sven Schwemmers, et al. "MPN patients harbor recurrent truncating mutations in transcription factor NF-E2." Journal of Experimental Medicine 210, no. 5 (April 15, 2013): 1003–19. http://dx.doi.org/10.1084/jem.20120521.
Full textAbstract:
The molecular etiology of myeloproliferative neoplasms (MPNs) remains incompletely understood, despite recent advances incurred through the discovery of several different mutations in MPN patients. We have recently described overexpression of the transcription factor NF-E2 in MPN patients and shown that elevated NF-E2 levels in vivo cause an MPN phenotype and predispose to leukemic transformation in transgenic mice. We report the presence of acquired insertion and deletion mutations in the NF-E2 gene in MPN patients. These result in truncated NF-E2 proteins that enhance wild-type (WT) NF-E2 function and cause erythrocytosis and thrombocytosis in a murine model. NF-E2 mutant cells acquire a proliferative advantage, witnessed by clonal dominance over WT NF-E2 cells in MPN patients. Our data underscore the role of increased NF-E2 activity in the pathophysiology of MPNs.
8
Mutschler, Manuel, Angela S. Magin, Martina Buerge, Britta Will, Ingo H. Pilz, Anna Rita Migliaccio, and Heike L. Pahl. "NF-E2 Overexpression Delays Erythroid Differentiation and Increases Erythrocyte Production." Blood 110, no. 11 (November 16, 2007): 1546. http://dx.doi.org/10.1182/blood.v110.11.1546.1546.
Full textAbstract:
Abstract The transcription factor Nuclear-Factor-Erythroid2 (NF-E2) is overexpressed in the vast majority of patients with polycythemia vera (PV). PV affects precisely those lineages expressing NF-E2: hematopoietic precursors, erythroid, megakaryocytic and granulocytic cells. In murine models, NF-E2 overexpression increases proliferation and promotes cellular viability in the absence of EPO. EPO-independent growth is a hallmark of PV. We therefore hypothesized that NF-E2 overexpression contributes to erythrocytosis, the pathognomonic symptom of PV. Consequently, we investigated the effect of NF-E2 overexpression in healthy CD34+ cells. NF-E2 overexpression lead to a delay in erythroid differentiation, manifested by a belated appearance of GlycophorinA-positive mature erythroid cells. Differentiation delay was similarly observed in primary PV patient erythroid cultures compared to healthy controls. Protracted differentiation lead to a significant increase in the accumulated number of erythroid cells both in PV cultures and in CD34+ cells overexpressing NF-E2. Similarly, NF-E2 overexpression altered erythroid colony formation, leading to an increase in BFU-E formation. These data argue that NF-E2 overexpression delays the early phase of erythroid differentiation, resulting in an expansion of erythroid progenitors, thereby increasing the number of erythrocytes derived from one CD34+ cell. These data propose a role for NF-E2 in mediating the erythrocytosis of PV.
9
Wang, Wei, Sven Schwemmers, and Heike L. Pahl. "AML-1/RUNX-1 Is Overexpressed in Patients with Myeloproliferative Neoplasms and Mediates JAK2 V617F-Independent Overexpression of NF-E2." Blood 114, no. 22 (November 20, 2009): 3893. http://dx.doi.org/10.1182/blood.v114.22.3893.3893.
Full textAbstract:
Abstract Abstract 3893 Poster Board III-829 The transcription factor nuclear factor erythroid-2 (NF-E2) plays an essential role in both erythropoiesis and megakaryopoiesis. In addition to erythroid, megakaryocytic and granulocytic progenitors, NF-E2 is expressed in hematopoietic stem cells and in mature granulocytes. We have previously shown that NF-E2 is overexpressed in the vast majority of patients with polycythemia vera (PV). Concomitantly, 90 – 95% of these patients carry the JAK2V617F point mutation. Even though NF-E2 levels correlated with JAK2V671F allele burden in some PV cohorts, the molecular mechanism causing aberrant NF-E2 expression in PV patients has not been described. Here we show that NF-E2 expression is also increased in patients with Essential Thrombocythemia (ET) and that this is independent of the presence of the JAK2V617F mutation. Characterization of the NF-E2 promoter revealed multiple binding sites for Acute Myeloid Leukemia-1 (AML-1/RUNX-1), which were shown to be functional by deletion analysis and site directed mutagenesis. Chromatin Immunoprecipitation (ChIP) demonstrated AML-1 binding to the NF-E2 promoter in vivo. Moreover, AML-1 binding to the NF-E2 promoter in vivo is significantly increased in primary granulocytes from PV patients compared to healthy controls. Subsequently, we demonstrated that mRNA and protein expression of AML-1 is elevated in PV and ET patients, both in the presence and absence of JAK2V617F. In addition, AML-1 and NF-E2 expression are highly correlated. RNAi-mediated suppression of AML-1 levels significantly decreased NF-E2 expression. In summary, our data identify NF-E2 as a novel AML-1 target gene and point to a role of aberrant AML-1 expression in mediating elevated NF-E2 expression in MPN patients. Disclosures: No relevant conflicts of interest to declare.
10
Kashif, Muhammed, Hellwig Andrea, Herzog Stefanie, Vinnikov Ilya, Shahzad Khurrum, Nawroth Peter, and Berend Isermann. "The Transcription Factor NF-E2 Regulates Trophoblast Differentiation and Placental Vascularisation Independent of Platelets." Blood 112, no. 11 (November 16, 2008): 3927. http://dx.doi.org/10.1182/blood.v112.11.3927.3927.
Full textAbstract:
Abstract Embryonic growth restriction is associated with increased perinatal morbidity and mortality. We previously established that the embryonic lack of the transcription factor NF-E2 results in intrauterine growth restriction (IUGR) associated with a reduced placental vascularisation. The mechanism underlying the reduced placental vascularisation in embryos lacking NF-E2 remains unknown. The transcription factor NF-E2 is expressed in cells of the erythrocyte and megakaryocyte lineage. Absence of NF-E2 impairs thrombocytopoiesis secondary to a megakaryocyte defect, resulting in severe thrombocytopenia. The mechanisms underlying the IUGR and reduced placental vascularisation, in particular the role of thrombocytopenia, remains unknown. Analysis of placental vascularisation and embryonic growth revealed a reduction of vascularisation in NF-E2−/− placenta as early as day 14.5 p.c., while the growth retardation of NF-E2−/− embryos was only observed at day 17.5 p.c. Therefore, the placental phenotype precedes embryonic growth retardation, consistent with a primary defect within the placenta. RT-PCR and in situ hybridization detected expression of NF-E2 in the labyrinthine layer and the junctional zone of the E 14.5 placenta as well as in murine trophoblast stem cells in vitro. To determine whether platelet deficiency or a trophoblast specific defect impairs placental vascularisation various in vivo experiments were conducted. Restoring platelet generation in NF-E2 knock out embryos using tetraploid aggregation or via in utero megakaryocyte transplantation does not rescue the vascularisation defect. Consistently, platelet depletion in NF-E2 expressing embryos is not sufficient to reduce placental vascularisation. Therefore, the impaired vascularisation in NF-E2 deficient placenta is independent of the platelet deficiency and results from NF-E2-deficieny in trophoblast cells. Expression analysis (RT-PCR, in situ hybridisation) showed up regulation of syncytiotrophoblast marker Gcm1 and down regulation of labyrinthine layer marker Esx1 in NF-E2−/− placentae. Using electron microscopy we detected thickening of labyrinth trophoblast layers (syncytiotrophoblast layer II and III). To further delineate the mechanisms gene-expression analyses was performed. A number of genes known to be relevant for placental development were less expressed in the absence of NF-E2. Among these genes Fra-1, a member of the AP-1 dimeric transcription factor family with an established role for placental labyrinthine formation, was markedly reduced. Using EMSA we detected a marked reduction of AP-1 binding activity in placental extracts of NF-E2 deficient embryos. These data establish that NF-E2 and Fra-1 interact directly, as has been previously established, or indirectly, through NF-E2 dependent regulation of Fra-1 expression in trophoblast cells, to regulate placental vascularisation. These experiments identify a role of the transcription factor NF-E2 in non-hematopoetic cells. NF-E2 regulates expression of genes relevant for placental development, including the transcription factor Fra-1. The absence of NF-E2 results in impaired placental vascularisation and enhanced syncytium formation independent of platelets through a trophoblast specific defect. These studies therefore identify a new NF-E2 dependent mechanism underlying IUGR.
11
Jutzi, Jonas S., Albert Gründer, Sandra Kaiser, Gorica Nikoloski, Joop H. Jansen, and Heike L. Pahl. "Mutations In Transcription Factor NF-E2 Confer Resistance To Interferon Treatment In MPN Patients." Blood 122, no. 21 (November 15, 2013): 4096. http://dx.doi.org/10.1182/blood.v122.21.4096.4096.
Full textAbstract:
Abstract Recently, we described insertion and deletion mutations in transcription factor NF-E2 in a subset of patients with Myeloproliferative Neoplasms (MPN), most frequently patients with primary or secondary myelofibrosis (PMF or sMF, respectively) as well as patients with polycythemia vera (PV). These mutations lead to the formation of truncated NF-E2 proteins that lack DNA binding activity. Nonetheless, mutant NF-E2 proteins augment wild-type NF-E2 activity both in vitro and in vivo in a murine bone marrow transplant model. Mice expressing mutant NF-E2 develop erythrocytosis and thrombocytosis. Here, we demonstrate that, in contrast to mice expressing wild-type NF-E2, treatment of mice expressing mutant NF-E2 with interferon-alpha does not lower red blood cell numbers (RBC), white blood cell counts (WBC) or platelet numbers. While these parameters are significantly decreased by interferon-alpha (IFN) in mice expressing wild-type NF-E2, mice expressing mutant NF-E2 showed no response to interferon treatment in these counts over a four month period. Two PV patients carrying truncating mutations in NF-E2 received IFN treatment. Blood samples were available both from before treatment as well as from 3 and 4 years after the beginning of therapy, respectively. Purified granulocytes from these samples were analyzed for mutant NF-E2 allele burden by GeneScan analysis. In both patients, the mutant NF-E2 allele burden rose during IFN treatment, increasing from 1% to 30% in one patient and from 24% to 100% in the second. We have demonstrated that acquisition of an NF-E2 mutation confers a proliferative advantage on cells already carrying the JAK2V617F mutation. Using colony assays from primary MPN cells, we showed that cells carrying both mutations outcompete cells carrying only the JAK2V617F mutation in vivo. Cells expressing both mutant NF-E2 and JAK2V617F are statistically significantly more frequently in the S-phase of the cell cycle and express significantly higher levels of several proteins that promote G1/S transition, cyclin D3, CDK4 and CDK6. The data presented here suggest that IFN treatment is insufficient to counteract the proliferative drive conferred by mutant NF-E2. Disclosures: No relevant conflicts of interest to declare.
12
Mignotte, V., J. F. Eleouet, N. Raich, and P. H. Romeo. "Cis- and trans-acting elements involved in the regulation of the erythroid promoter of the human porphobilinogen deaminase gene." Proceedings of the National Academy of Sciences 86, no. 17 (September 1989): 6548–52. http://dx.doi.org/10.1073/pnas.86.17.6548.
Full textAbstract:
Two cis-acting sequences, recognized by two erythroid-specific trans-acting factors, are involved in the regulation of the erythroid promoter of the human gene coding for porphobilinogen deaminase (PBGD). The first region, located at -70, binds the erythroid factor NF-E1, and point mutations within this region abolish the induction of transcription of this promoter during murine erythroleukemia (MEL) cell differentiation. The second region, located at -160, binds the erythroid-specific factor NF-E2 and the ubiquitous factor AP1. Using UV cross-linking, we show that NF-E2 has a higher molecular weight than AP1, demonstrating that NF-E2 is not an erythroid-specific degradation product of AP1. By point mutagenesis of the NF-E2/AP1 binding site, we define mutations that abolish binding of either NF-E2 alone or AP1 and NF-E2 together. Regulation of transcription of the PBGD erythroid promoter is abolished by those mutations, suggesting that NF-E2 but not AP1 is necessary for correct regulation of this promoter in erythroid cells.
13
Koellerer, Christoph, Heike L. Pahl, and Julius Wehrle. "The Cell Cycle Regulators CDK4, CDK6 and Cyclin D3 As Well As the Histone Methyltransferases MLL2 and MLL4 Constitute NF-E2 Target Genes That Participate in a Novel Regulatory Loop." Blood 124, no. 21 (December 6, 2014): 4592. http://dx.doi.org/10.1182/blood.v124.21.4592.4592.
Full textAbstract:
Abstract Background: Expression of the transcription factor and epigenetic modulator “nuclear factor erythroid-2” (NF-E2) is aberrantly elevated in patients with Myeloproliferative Neoplasms (MPN). We have shown that NF-E2 overexpression in a murine model causes a MPN phenotype. In addition, we have identified in-del mutations in the NF-E2 gene in MPN patients, which, despite having lost the ability to bind DNA, nonetheless increase the activity of wild type NF-E2. These mutants, which are acquired secondary to the JAK2V617F mutation, confer a proliferative advantage beyond that provided by the JAK2V617F alone. In all patients analyzed, cells carrying both mutations displayed clonal dominance, vastly outnumbering those carrying JAK2V617Falone. Mutant NF-E2 causes an increase in the number of cells entering S-phase mediated by increased expression of the cell cycle regulators CDK4, CDK6 and CyclinD3 that promote the G1-S transition. However, it is not known whether these cell cycle regulators constitute direct NF-E2 target genes or whether the proliferative effect is indirect. Moreover, additional downstream pathways by which this augmented transcription factor activity exerts its effects remain unknown. Aims: We therefore investigated whether CDK4, CDK6 and Cyclin D3 constitute direct NF-E2 target genes. Moreover, we identified additional novel NF-E2 targets in an effort to delineate the molecular mechanism by which increased NF-E2 activity contributes to MPN pathophysiology. Methods: We used ChIP and luciferase reporter assays as well as qRT-PCR and Western Blot to identify and characterize novel NF-E2 targets. Results: Our data show that both the cell cycle regulators CDK4, CDK6 and CyclinD3 as well as the histone methyltransferases MLL2 and MLL4 constitute direct NF-E2 target genes, as the transcription factor binds these genes in vivo. ChIP analysis of HEL cells shows NF-E2 binding to a site in the CDK4 promoter, four sites in the CDK6 and the Cyclin D3 locus each, to the MLL4 promoter as well as to three sites in the MLL2 locus, establishing these five genes as direct NF-E2 targets. Activity of NF-E2 on the MLL4 promoter was confirmed by reporter gene assays. Expression of both the cell cycle regulators and the methyltransferases was statistically significantly elevated in bone marrow from NF-E2 overexpressing mice. Most importantly, quantitative RT-PCR showed significantly increased mRNA expression of all five target genes in primary cells of patients with polycythemia vera (PV) compared to healthy controls. Hence, the expression of the cell cycle regulators and the epigenetically active enzymes is augmented in PV patients. Corresponding to the increased methyltransferase activity, primary cells from PV patients showed a significant elevation in global H3K4m1 levels, a chromatin mark conferred by the MLL proteins. The MLL2 protein has been previously shown to interact with NF-E2. NF-E2 serves as a scaffold and is required both for recruitment of MLL2 to the beta-globin locus as well as for chromatin remodeling at these sites. Here we show that MLL2 binds the CDK6 locus at the same sites as NF-E2, strongly suggesting that NF-E2 likewise acts to direct MLL2 to the CDK6 gene. Our data establish a novel interaction network where NF-E2 both regulates the expression levels of MLL2 while at the same time modulating its epigenetic activity at NF-E2 target genes, which include critical cell cycle regulators. Summary / Conclusions: Here we show that the cell cycle regulators CDK4, CDK6 and Cyclin D3 as well as the histone methyltransferases MLL2 and MLL4 constitute novel NF-E2 target genes. We propose that elevated NF-E2 levels in MPN patients contribute to disease pathophysiology by participating in a novel regulatory loop, which both enhances transcription and alteres histone methylation leading to increased proliferation. These data provide a molecular basis for pre-clinical investigation into the effects of CDK 4/6 inhibitors as well as of histone methyltransferase inhibitors on MPN cell biology. Disclosures No relevant conflicts of interest to declare.
14
Peeken, Jan, Thalia S. Seeger, Julius Wehrle, Daniel H. Schanne, Monika Gothwal, Albert Gründer, and Heike L. Pahl. "Overexpression Of The Histone Demethylase JMJD1C In Polycythemia Vera Contributes To NF-E2 Overexpression Via Epigenetic Dysregulation and An Auto-Regulatory Loop." Blood 122, no. 21 (November 15, 2013): 1602. http://dx.doi.org/10.1182/blood.v122.21.1602.1602.
Full textAbstract:
Abstract Expression of the transcription factor and epigenetic modulator “nuclear factor erythroid-2” (NF-E2) is aberrantly elevated in patients with Myeloproliferative Neoplasms (MPN). We have recently shown that NF-E2 overexpression in a murine model causes a phenotype similar to MPN in humans. This includes thrombocytosis, leukocytosis, expansion of the stem- and progenitor cell compartments as well as spontaneous transformation to acute leukemia. However, both the downstream pathways by which this transcription factor exerts its effects as well as the mechanisms leading to its overexpression in MPN patients remain incompletely characterized. We show here that the histone demethylases JMJD1C and JMJD2C constitute novel NF-E2 target genes. Using chromatin immunoprecipitation (ChIP) we demonstrate NF-E2 binding to the JMJD1C and JMJD2C loci. Both JMJD1C and JMJD2C protein levels are statistically significantly elevated in patients with polycythemia vera (PV) compared to healthy controls. JMJD1C and 2C proteins mediate the demethylation of histone H3K9, converting H3K9me2 via H3K9me to unmethylated H3K9. Consistent with increased demethylase activity, we observed statistically significantly decreased levels of global H3K9me and H3K9me2 in PV patients compared to healthy controls. We subsequently investigated whether the NF-E2 gene is regulated by epigenetic mechanisms and whether these are altered in PV patients. Indeed, using ChIP we demonstrate presence of the repressive H3K9me2 mark at three sites within the NF-E2 locus in healthy control granulocytes, where NF-E2 is expressed at low levels. These repressive histone marks are significantly reduced or completely absent in PV patients, which overexpress NF-E2. At the same time, binding of the repressive heterochromatin protein 1-alpha (HP1a) to the NF-E2 gene, while present in healthy controls, is significantly reduced in PV patients. HP1a chromatin binding is dependent on the presence of the H3K9me2 modification. Decreased levels of H3K9me2 and decreased HP1a binding are therefore both consistent with elevated NF-E2 expression in PV. We hypothesized that increased levels of the H3K9me2 demethylase JMJD1C in PV patients are responsible for the loss of this histone mark in the NF-E2 gene. Consistent with this model, we demonstrate significantly increased binding of JMJD1C to the NF-E2 locus in purified primary PV granulocytes. Our data demonstrate that the NF-E2 gene is regulated by epigenetic mechanisms and that these histone modifications are perturbed in PV patients. Moreover, NF-E2 participates in an auto-regulatory loop, by directly regulating transcription of the histone demethylase JMJD1C. Increased levels of NF-E2 thus cause increased JMJD1C expression. The JMJD1C protein, in turn, demethylates H3K9me2 residues in the NF-E2 locus, thereby further augmenting NF-E2 expression. Besides sustaining this novel autoregulatory loop, increased JMJD1C activity may contribute to MPN pathology by altering gene expression at additional loci. We sought to determine whether elevated NF-E2 levels could be normalized by pharmacological intervention. Treatment with the nucleoside analogue Decitabine reduced NF-E2 expression, and restored both the H3K9me2 marks and HP1a binding to the NF-E2 locus. These data provide a molecular rational for pre-clinical investigation of the effects of histone demethylase inhibitors and Decitabine on MPN cell biology and, subsequently, for a phase I trial investigating a combination of Decitabine and histone demethylase inhibitors in MPN patients. Disclosures: No relevant conflicts of interest to declare.
15
Cheng, X., M. J. Reginato, N. C. Andrews, and M. A. Lazar. "The transcriptional integrator CREB-binding protein mediates positive cross talk between nuclear hormone receptors and the hematopoietic bZip protein p45/NF-E2." Molecular and Cellular Biology 17, no. 3 (March 1997): 1407–16. http://dx.doi.org/10.1128/mcb.17.3.1407.
Full textAbstract:
Thyroid hormone (T3) and retinoic acid (RA) play important roles in erythropoiesis. We found that the hematopoietic cell-specific bZip protein p45/NF-E2 interacts with T3 receptor (TR) and RA receptor (RAR) but not retinoid X receptor. The interaction is between the DNA-binding domain of the nuclear receptor and the leucine zipper region of p45/NF-E2 but is markedly enhanced by cognate ligand. Remarkably, ligand-dependent transactivation by TR and RAR is markedly potentiated by p45/NF-E2. This effect of p45/NF-E2 is prevented by maf-like protein p18, which functions positively as a heterodimer with p45/NF-E2 on DNA. Potentiation of hormone action by p45/NF-E2 requires its activation domain, which interacts strongly with the multifaceted coactivator cyclic AMP response element protein-binding protein (CBP). The region of CBP which interacts with p45/NF-E2 is the same interaction domain that mediates inhibition of hormone-stimulated transcription by AP1 transcription factors. Overexpression of the bZip interaction domain of CBP specifically abolishes the positive cross talk between TR and p45/NF-E2. Thus, positive cross talk between p45/NF-E2 and nuclear hormone receptors requires direct protein-protein interactions between these factors and with CBP, whose integration of positive signals from two transactivation domains provides a novel mechanism for potentiation of hormone action in hematopoietic cells.
16
Levin, Jack, Jin-Peng Peng, Georgiann R. Baker, Jean-Luc Villeval, Patrick Lecine, Samuel A. Burstein, and Ramesh A. Shivdasani. "Pathophysiology of Thrombocytopenia and Anemia in Mice Lacking Transcription Factor NF-E2." Blood 94, no. 9 (November 1, 1999): 3037–47. http://dx.doi.org/10.1182/blood.v94.9.3037.
Full textAbstract:
Abstract Expression of the p45 subunit of transcription factor NF-E2 is restricted to selected blood cell lineages, including megakaryocytes and developing erythrocytes. Mice lacking p45 NF-E2 show profound thrombocytopenia, resulting from a late arrest in megakaryocyte differentiation, and a number of red blood cell defects, including anisocytosis and hypochromia. Here we report results of studies aimed to explore the pathophysiology of these abnormalities. Mice lacking NF-E2 produce very few platelet-like particles that display highly disorganized ultrastructure and respond poorly to platelet agonists, features consistent with the usually lethal hemorrhage in these animals. Thrombocytopenia was evident during fetal life and was not corrected by splenectomy in adults. Surprisingly, fetal NF-E2–deficient megakaryocyte progenitors showed reduced proliferation potential in vitro. Thus, NF-E2 is required for regulated megakaryocyte growth as well as for differentiation into platelets. All the erythroid abnormalities were reproduced in lethally irradiated wild-type recipients of hematopoietic cells derived from NF-E2-null fetuses. Whole blood from mice lacking p45 NF-E2 showed numerous small red blood cell fragments; however, survival of intact erythrocytes in vivo was indistinguishable from control mice. Considered together, these observations indicate a requirement for NF-E2 in generating normal erythrocytes. Despite impressive splenomegaly at baseline, mice lacking p45 NF-E2 survived splenectomy, which resulted in increased reticulocyte numbers. This reveals considerable erythroid reserve within extra-splenic sites of hematopoiesis and suggests a role for the spleen in clearing abnormal erythrocytes. Our findings address distinct aspects of the requirements for NF-E2 in blood cell homeostasis and establish its roles in proper differentiation of megakaryocytes and erythrocytes.
17
Levin, Jack, Jin-Peng Peng, Georgiann R. Baker, Jean-Luc Villeval, Patrick Lecine, Samuel A. Burstein, and Ramesh A. Shivdasani. "Pathophysiology of Thrombocytopenia and Anemia in Mice Lacking Transcription Factor NF-E2." Blood 94, no. 9 (November 1, 1999): 3037–47. http://dx.doi.org/10.1182/blood.v94.9.3037.421k42_3037_3047.
Full textAbstract:
Expression of the p45 subunit of transcription factor NF-E2 is restricted to selected blood cell lineages, including megakaryocytes and developing erythrocytes. Mice lacking p45 NF-E2 show profound thrombocytopenia, resulting from a late arrest in megakaryocyte differentiation, and a number of red blood cell defects, including anisocytosis and hypochromia. Here we report results of studies aimed to explore the pathophysiology of these abnormalities. Mice lacking NF-E2 produce very few platelet-like particles that display highly disorganized ultrastructure and respond poorly to platelet agonists, features consistent with the usually lethal hemorrhage in these animals. Thrombocytopenia was evident during fetal life and was not corrected by splenectomy in adults. Surprisingly, fetal NF-E2–deficient megakaryocyte progenitors showed reduced proliferation potential in vitro. Thus, NF-E2 is required for regulated megakaryocyte growth as well as for differentiation into platelets. All the erythroid abnormalities were reproduced in lethally irradiated wild-type recipients of hematopoietic cells derived from NF-E2-null fetuses. Whole blood from mice lacking p45 NF-E2 showed numerous small red blood cell fragments; however, survival of intact erythrocytes in vivo was indistinguishable from control mice. Considered together, these observations indicate a requirement for NF-E2 in generating normal erythrocytes. Despite impressive splenomegaly at baseline, mice lacking p45 NF-E2 survived splenectomy, which resulted in increased reticulocyte numbers. This reveals considerable erythroid reserve within extra-splenic sites of hematopoiesis and suggests a role for the spleen in clearing abnormal erythrocytes. Our findings address distinct aspects of the requirements for NF-E2 in blood cell homeostasis and establish its roles in proper differentiation of megakaryocytes and erythrocytes.
18
Forsberg, E. Camilla, Karen M. Downs, and Emery H. Bresnick. "Direct interaction of NF-E2 with hypersensitive site 2 of the β-globin locus control region in living cells." Blood 96, no. 1 (July 1, 2000): 334–39. http://dx.doi.org/10.1182/blood.v96.1.334.
Full textAbstract:
Abstract The human β-globin locus control region (LCR) confers high-level, tissue-specific expression to the β-globin genes. Tandem Maf recognition elements (MAREs) within the hypersensitive site 2 (HS2) subregion of the LCR are important for the strong enhancer activity of the LCR. Multiple proteins are capable of interacting with these sites in vitro, including the erythroid cell- and megakaryocyte-specific transcription factor, NF-E2. The importance of NF-E2 for β-globin gene expression is evident in murine erythroleukemia cells lacking the p45 subunit of NF-E2. These CB3 cells have a severe defect in - and β-globin gene transcription, which can be restored by expression of NF-E2. However, mice nullizygous for p45 express nearly normal levels of β-globin. Thus, either a redundant factor(s) exists in mice that can functionally replace NF-E2, or NF-E2 does not function through the LCR to regulate β-globin gene expression. To address this issue, we asked whether NF-E2 binds directly to the tandem MAREs of HS2 in intact cells. Using a chromatin immunoprecipitation assay, we provide evidence for NF-E2 binding directly and specifically to HS2 in living erythroleukemia cells and in mouse fetal liver. The specific immunoisolation of HS2 sequences was dependent on the presence of p45 and on intact MAREs within HS2. These results support a direct role for NF-E2 in the regulation of β-globin gene expression through activation of the LCR.
19
Forsberg, E. Camilla, Karen M. Downs, and Emery H. Bresnick. "Direct interaction of NF-E2 with hypersensitive site 2 of the β-globin locus control region in living cells." Blood 96, no. 1 (July 1, 2000): 334–39. http://dx.doi.org/10.1182/blood.v96.1.334.013k17_334_339.
Full textAbstract:
The human β-globin locus control region (LCR) confers high-level, tissue-specific expression to the β-globin genes. Tandem Maf recognition elements (MAREs) within the hypersensitive site 2 (HS2) subregion of the LCR are important for the strong enhancer activity of the LCR. Multiple proteins are capable of interacting with these sites in vitro, including the erythroid cell- and megakaryocyte-specific transcription factor, NF-E2. The importance of NF-E2 for β-globin gene expression is evident in murine erythroleukemia cells lacking the p45 subunit of NF-E2. These CB3 cells have a severe defect in - and β-globin gene transcription, which can be restored by expression of NF-E2. However, mice nullizygous for p45 express nearly normal levels of β-globin. Thus, either a redundant factor(s) exists in mice that can functionally replace NF-E2, or NF-E2 does not function through the LCR to regulate β-globin gene expression. To address this issue, we asked whether NF-E2 binds directly to the tandem MAREs of HS2 in intact cells. Using a chromatin immunoprecipitation assay, we provide evidence for NF-E2 binding directly and specifically to HS2 in living erythroleukemia cells and in mouse fetal liver. The specific immunoisolation of HS2 sequences was dependent on the presence of p45 and on intact MAREs within HS2. These results support a direct role for NF-E2 in the regulation of β-globin gene expression through activation of the LCR.
20
Kaufmann, Kai B., Albert Gründer, Tobias Hadlich, Julius Wehrle, Monika Gothwal, Ruzhica Bogeska, Thalia S. Seeger, et al. "A novel murine model of myeloproliferative disorders generated by overexpression of the transcription factor NF-E2." Journal of Experimental Medicine 209, no. 1 (January 9, 2012): 35–50. http://dx.doi.org/10.1084/jem.20110540.
Full textAbstract:
The molecular pathophysiology of myeloproliferative neoplasms (MPNs) remains poorly understood. Based on the observation that the transcription factor NF-E2 is often overexpressed in MPN patients, independent of the presence of other molecular aberrations, we generated mice expressing an NF-E2 transgene in hematopoietic cells. These mice exhibit many features of MPNs, including thrombocytosis, leukocytosis, Epo-independent colony formation, characteristic bone marrow histology, expansion of stem and progenitor compartments, and spontaneous transformation to acute myeloid leukemia. The MPN phenotype is transplantable to secondary recipient mice. NF-E2 can alter histone modifications, and NF-E2 transgenic mice show hypoacetylation of histone H3. Treatment of mice with the histone deacetylase inhibitor (HDAC-I) vorinostat restored physiological levels of histone H3 acetylation, decreased NF-E2 expression, and normalized platelet numbers. Similarly, MPN patients treated with an HDAC-I exhibited a decrease in NF-E2 expression. These data establish a role for NF-E2 in the pathophysiology of MPNs and provide a molecular rationale for investigating epigenetic alterations as novel targets for rationally designed MPN therapies.
21
Jutzi, Jonas S., A. Gruender, Konrad Aumann, and Heike L. Pahl. "ITCH Deficiency Promotes an MPN Phenotype with Eosinophilia, Inflammation, Mislocalization and Overexpression of the Transcription Factor NF-E2." Blood 124, no. 21 (December 6, 2014): 821. http://dx.doi.org/10.1182/blood.v124.21.821.821.
Full textAbstract:
Abstract Background: We have described overexpression of the transcription factor NF-E2 in MPN patients and shown that elevated NF-E2 levels cause a MPN phenotype in transgenic mice. This includes thrombocytosis, leukocytosis, splenomegaly as well as an expansion of the stem- and progenitor cell compartments in the bone marrow. Recently, we have shown that, counterintuitively for a transcription factor, NF-E2 is located exclusively in the cytoplasm in the vast majority of erythroid cells in the bone marrow (85%). Patients with PMF show a statistically highly significant elevation in the proportion of cells displaying nuclear NF-E2 compared to either healthy controls or ET and PV patients. However, the molecular mechanisms regulating the subcellular localization of NF-E2 and its aberrant localization in PMF remain to be investigated. The E3 ubiquitin ligase ITCH has been postulated to stabilize and retain NF-E2 in the cytosol by protein-protein interaction and subsequent ubiquitinylation. The phenotype of ITCH deficient mice, however, has only been described briefly: animals display splenomegaly and an expansion of the stem cell compartment. The effect of ITCH deficiency on peripheral blood counts and on NF-E2 activity has not been determined. Aims: To characterize the phenotype of ITCH deficient mice and investigate the effect of ITCH deficiency on NF-E2 localization and activity. Methods: The peripheral blood and bone marrow of ITCH knock out mice as well as of heterozygous and wild-type control animals was analyzed: CBCs were determined every four weeks, stem- and progenitor populations in the bone marrow were assessed by 7-color FACS. Expression levels of NF-E2 and its targets genes were measured by quantitative PCR. Plasma cytokine concentrations were measured by Cytometric Bead Array. To determine the subcellular localization of NF-E2, immunohistochemical stainings of ITCH knock out BMs and wild-type controls were conducted. Results: At several consecutive time points ITCH knock out mice displayed a statistically significant elevation in WBC compared to heterozygous and wild-type littermates. Interestingly, both the percentage and the absolute number of eosinophils were significantly increased, some animals presenting with a drastic eosinophilia, the differential containing over 60% eosinophils. Furthermore, ITCH knock out mice display a significant decrease in platelet count, accompanied by an increase in platelet mass and volume, indicative of giant platelets. In the bone marrow ITCH deficient mice show a significant increase in the absolute number of Common Myeloid Progenitors (CMP). NF-E2 expression levels in the peripheral blood as well as in the bone marrow were highly statistically significantly increased compared to the levels measured in wild-type or heterozygous control mice. Consequently, the NF-E2 target gene Thromboxane Synthase A was statistically significantly overexpressed in peripheral blood of ITCH knock out mice. Plamsa concentrations of the inflammatory cytokines INF-γ and TNF were statistically significantly elevated, reaching two to threefold higher levels in ITCH knock out mice compared to wild-type littermates. Lastly, NF-E2 subcellular localization was altered in ITCH deficient mice, which display a significant increase in the proportion of megakaryocytes positive for nuclear NF-E2. Summary/Conclusions: Our data identify the E3 ubiquitin ligase ITCH as a regulator of NF-E2 activity. Impaired ITCH activity leads to both an NF-E2 overexpression and an increased nuclear NF-E2 localization that together drive overexpression of NF-E2 target genes. Furthermore, ITCH deficiency leads to higher inflammatory cytokine levels, comparable to those seen in PMF patients. All of these factors contribute to the resulting myeloproliferative phenotype with eosinophilia. Our data provide the first pathophysiological explanation of the pathognomonic symptom of ITCH deletion: pruritus in "itchy" mice. Moreover, given the aberrant NF-E2 localization in PMF patients, our data provide a possible mechanism and underscore the role of elevated NF-E2 activity in the pathophysiology of myeloproliferative neoplasms. Disclosures No relevant conflicts of interest to declare.
22
Shyu, Yu-Chiau, Tung-Liang Lee, Chun-Yuan Ting, Shau-Ching Wen, Lie-Jiau Hsieh, Yueh-Chun Li, Jau-lang Hwang, Chyi-Chyang Lin, and C. K. James Shen. "Sumoylation of p45/NF-E2: Nuclear Positioning and Transcriptional Activation of the Mammalian β-Like Globin Gene Locus." Molecular and Cellular Biology 25, no. 23 (December 1, 2005): 10365–78. http://dx.doi.org/10.1128/mcb.25.23.10365-10378.2005.
Full textAbstract:
ABSTRACT NF-E2 is a transcription activator for the regulation of a number of erythroid- and megakaryocytic lineage-specific genes. Here we present evidence that the large subunit of mammalian NF-E2, p45, is sumoylated in vivo in human erythroid K562 cells and in mouse fetal liver. By in vitro sumoylation reaction and DNA transfection experiments, we show that the sumoylation occurs at lysine 368 (K368) of human p45/NF-E2. Furthermore, p45 sumoylation enhances the transactivation capability of NF-E2, and this is accompanied by an increase of the NF-E2 DNA binding affinity. More interestingly, we have found that in K562 cells, the β-globin gene loci in the euchromatin regions are predominantly colocalized with the nuclear bodies promyelocytic leukemia protein (PML) oncogenic domains that are enriched with the PML, SUMO-1, RNA polymerase II, and sumoylatable p45/NF-E2. Chromatin immunoprecipitation assays further showed that the intact sumoylation site of p45/NF-E2 is required for its binding to the DNase I-hypersensitive sites of the β-globin locus control region. Finally, we demonstrated by stable transfection assay that only the wild-type p45, but not its mutant form p45 (K368R), could efficiently rescue β-globin gene expression in the p45-null, erythroid cell line CB3. These data together point to a model of mammalian β-like globin gene activation by sumoylated p45/NF-E2 in erythroid cells.
23
Blank, Volker, Min J. Kim, and Nancy C. Andrews. "Human MafG Is a Functional Partner for p45 NF-E2 in Activating Globin Gene Expression." Blood 89, no. 11 (June 1, 1997): 3925–35. http://dx.doi.org/10.1182/blood.v89.11.3925.
Full textAbstract:
Abstract Mammalian globin gene expression is activated through NF-E2 elements recognized by basic-leucine zipper proteins of the AP-1 superfamily. The specificity of NF-E2 DNA binding is determined by several nucleotides adjacent to a core AP-1 motif, comprising a recognition site for transcription factors of the Maf subfamily. Earlier work proposed that p18(MafK) forms a heterodimer with hematopoietic-specific protein p45 NF-E2 to activate transcription through NF-E2 sites. However, there was no direct evidence that p18(MafK) serves this function in vivo; in fact, mice lacking p18(MafK) have no phenotype. Here we describe a novel cDNA clone that encodes the human homolog of chicken MafG. Human MafG heterodimerizes with p45 NF-E2 and binds DNA with specificity identical to that of purified NF-E2 DNA binding activity. A tethered heterodimer of p45 and MafG is fully functional in supporting expression of α- and β-globin, and in promoting erythroid differentiation in CB3, a p45-deficient mouse erythroleukemia cell line. These results indicate that human MafG can serve as a functional partner for p45 NF-E2, and suggest that the p45/MafG heterodimer plays a role in the regulation of erythropoiesis.
24
Gruender, Albert, Kai Kaufmann, Tobias Hadlich, Thomas Günther, Roland Schüle, Annette H. Schmitt-Graeff, Attilio Orazi, and Heike L. Pahl. "Overexpression of NF-E2 In Vivo Causes Thrombocytosis and Acute Leukemia: A Murine Model of Myeloproliferative Neoplasms." Blood 114, no. 22 (November 20, 2009): 961. http://dx.doi.org/10.1182/blood.v114.22.961.961.
Full textAbstract:
Abstract Abstract 961 The transcription factor nuclear factor erythroid-2 (NF-E2) is expressed in hematopoietic stem cells as well as in myeloid, erythroid and megakaryocytic precursors. NF-E2 deficient mice display marked anemia at birth and die perinatally due to thrombopenia, demonstrating an essential role for NF-E2 in both in erythropoiesis and platelet formation. We have previously shown that NF-E2 is overexpressed in the vast majority of patients with Myeloproliferative Neoplasms (MPNs). However, the effect of augmented transcription factor activity has not been studied in vivo. We therefore engineered two independent transgenic mouse lines expressing human NF-E2 under the control of the vav-Promoter, which has previously been shown to direct transgene expression in hematopoietic stem cells as well as in precursor cells of all lineages. The two founder lines differed in the degree of NF-E2 overexpression displayed. While one line showed moderate overexpression (2 – 5-fold), the other line expressed human NF-E2 between 10 and 100-fold above the murine counterpart. Both lines paralleled observations in PV patients, where a wide range of NF-E2 overexpression was noted (median overexpression, 7-fold; range 2-fold to 40-fold; n = 59). The two founder lines show overlapping but distinct phenotypes. In both strains. moderately overexpressing NF-E2 transgenic mice (2 – 10-fold) invariably develop thrombocytosis with a latency of 14 months. In addition, megakaryocyte colony formation in the bone marrow is drastically increased. In contrast, thrombocytosis is not observed in the markedly overexpressing NF-E2 transgenic mice (above 20-fold). A similar inverse correlation between the degree of NF-E2 overexpression and platelet numbers was observed in MPN patients. In both strains, Epo-independent colony formation, a pathognomonic feature of polycythemia vera, is significantly increased in NF-E2 transgenic animals. Bone marrow histopathology shows findings characteristically seen in MPNs, including the presence of increased megakaryopoiesis with cytologically abnormal forms, often in clusters. Both NF-E2 transgenic strains display significantly increased mortality. Upon autopsy, between 15 and 20% of mice in both strains present with major gastrointestinal bleeding in conjunction with splenic atrophy. Spleen weight is reduced by over 50% (Transgenic mice: 49 +/-15 mg, wild type littermates 103 +/- 30 mg; p < 0.001, n = 8 each). One third of the remaining mice show moderate to marked splenomegaly (2 – 27 fold increase in spleen weight; mean: 434 mg, range: 124 – 2700 mg; p < 0.001 vs. wt littermates, n = 12). Histopathological examination of all spleens revealed mild to moderately expanded red pulp with increased numbers of iron containing histiocytes. This observation indicates increased red cell destruction and may explain the fact that neither hematocrit nor hemoglobin are elevated in NF-E2 transgenic animals. At 18 months of age, one mouse developed acute leukemia, which is currently being phenotyped. In summary, in a murine model moderate NF-E2 overexpression causes a phenotype resembling Essential Thrombocythemia. In addition, our preliminary data indicate that NF-E2 overexpression may predispose to the development of acute leukemia. Disclosures: No relevant conflicts of interest to declare.
25
Chan, J. Y., X. L. Han, and Y. W. Kan. "Isolation of cDNA encoding the human NF-E2 protein." Proceedings of the National Academy of Sciences 90, no. 23 (December 1, 1993): 11366–70. http://dx.doi.org/10.1073/pnas.90.23.11366.
Full textAbstract:
The human homolog of mouse NF-E2 was isolated from the K562 cell line and found to encode a member of the basic leucine-zipper family of DNA-binding regulatory proteins. The deduced amino acid sequence of the mouse and human proteins exhibited near identity. Comparison to the related protein, Nrf1, revealed significant homologies at isolated regions, particularly within the basic domain, suggesting that NF-E2 and Nrf1 are members of a distinct subfamily of basic leucine-zipper proteins that share similar DNA-binding properties. High levels of human NF-E2 mRNA were observed in human erythroleukemic cell lines examined. Extensive survey of human tissue samples found NF-E2 expression not limited to erythropoeitic organs. Expression in the colon and testis suggests that NF-E2 may participate in the regulation of genes other than globin.
26
Chan, J. Y., X. L. Han, and Y. W. Kan. "Cloning of Nrf1, an NF-E2-related transcription factor, by genetic selection in yeast." Proceedings of the National Academy of Sciences 90, no. 23 (December 1, 1993): 11371–75. http://dx.doi.org/10.1073/pnas.90.23.11371.
Full textAbstract:
We have devised a complementation assay in yeast to clone mammalian transcriptional activators and have used it to identify a human basic leucine-zipper transcription factor that we have designated Nrf1 for NF-E2-related factor 1. Nrf1 potentially encodes a 742-aa protein and displays marked homology to the mouse and human NF-E2 transcription factors. Nrf1 activates transcription via NF-E2 binding sites in yeast cells. The ubiquitous expression pattern of Nrf1 and the range of promoters containing the NF-E2 binding motif suggest that this gene may play a role in the regulation of heme synthesis and ferritin genes.
27
Armstrong, J. A., and B. M. Emerson. "NF-E2 disrupts chromatin structure at human beta-globin locus control region hypersensitive site 2 in vitro." Molecular and Cellular Biology 16, no. 10 (October 1996): 5634–44. http://dx.doi.org/10.1128/mcb.16.10.5634.
Full textAbstract:
The human beta-globin locus control region (LCR) is responsible for forming an active chromatin structure extending over the 100-kb locus, allowing expression of the beta-globin gene family. The LCR consists of four erythroid-cell-specific DNase I hypersensitive sites (HS1 to -4). DNase I hypersensitive sites are thought to represent nucleosome-free regions of DNA which are bound by trans-acting factors. Of the four hypersensitive sites only HS2 acts as a transcriptional enhancer. In this study, we examine the binding of an erythroid protein to its site within HS2 in chromatin in vitro. NF-E2 is a transcriptional activator consisting of two subunits, the hematopoietic cell-specific p45 and the ubiquitous DNA-binding subunit, p18. NF-E2 binds two tandem AP1-like sites in HS2 which form the core of its enhancer activity. In this study, we show that when bound to in vitro-reconstituted chromatin, NF-E2 forms a DNase I hypersensitive site at HS2 similar to the site observed in vivo. Moreover, NF-E2 binding in vitro results in a disruption of nucleosome structure which can be detected 200 bp away. Although NF-E2 can disrupt nucleosomes when added to preformed chromatin, the disruption is more pronounced when NF-E2 is added to DNA prior to chromatin assembly. Interestingly, the hematopoietic cell-specific subunit, p45, is necessary for binding to chromatin but not to naked DNA. Interaction of NF-E2 with its site in chromatin-reconstituted HS2 allows a second erythroid factor, GATA-1, to bind its nearby sites. Lastly, nucleosome disruption by NF-E2 is an ATP-dependent process, suggesting the involvement of energy-dependent nucleosome remodeling factors.
28
Lecine, Patrick, Joseph E. Italiano, Sang-We Kim, Jean-Luc Villeval, and Ramesh A. Shivdasani. "Hematopoietic-specific β1 tubulin participates in a pathway of platelet biogenesis dependent on the transcription factor NF-E2." Blood 96, no. 4 (August 15, 2000): 1366–73. http://dx.doi.org/10.1182/blood.v96.4.1366.
Full textAbstract:
Abstract The cellular and molecular bases of platelet release by terminally differentiated megakaryocytes represent important questions in cell biology and hematopoiesis. Mice lacking the transcription factor NF-E2 show profound thrombocytopenia, and their megakaryocytes fail to produce proplatelets, the microtubule-based precursors of blood platelets. Using mRNA subtraction between normal and NF-E2–deficient megakaryocytes, cDNA was isolated encoding β1 tubulin, the most divergent β tubulin isoform. In NF-E2–deficient megakaryocytes, β1 tubulin mRNA and protein are virtually absent. The expression of β1 tubulin is exquisitely restricted to platelets and megakaryocytes, where it appears late in differentiation and localizes to microtubule shafts and coils within proplatelets. Restoring NF-E2 activity in a megakaryoblastic cell line or in NF-E2–deficient primary megakaryocytes rescues the expression of β1 tubulin. Re-expressing β1 tubulin in isolation does not, however, restore proplatelet formation in the defective megakaryocytes, indicating that other critical factors are required; indeed, other genes identified by mRNA subtraction also encode structural and regulatory components of the cytoskeleton. These findings provide critical mechanistic links between NF-E2, platelet formation, and selected microtubule proteins, and they also provide novel molecular insights into thrombopoiesis.
29
Lecine, Patrick, Joseph E. Italiano, Sang-We Kim, Jean-Luc Villeval, and Ramesh A. Shivdasani. "Hematopoietic-specific β1 tubulin participates in a pathway of platelet biogenesis dependent on the transcription factor NF-E2." Blood 96, no. 4 (August 15, 2000): 1366–73. http://dx.doi.org/10.1182/blood.v96.4.1366.h8001366_1366_1373.
Full textAbstract:
The cellular and molecular bases of platelet release by terminally differentiated megakaryocytes represent important questions in cell biology and hematopoiesis. Mice lacking the transcription factor NF-E2 show profound thrombocytopenia, and their megakaryocytes fail to produce proplatelets, the microtubule-based precursors of blood platelets. Using mRNA subtraction between normal and NF-E2–deficient megakaryocytes, cDNA was isolated encoding β1 tubulin, the most divergent β tubulin isoform. In NF-E2–deficient megakaryocytes, β1 tubulin mRNA and protein are virtually absent. The expression of β1 tubulin is exquisitely restricted to platelets and megakaryocytes, where it appears late in differentiation and localizes to microtubule shafts and coils within proplatelets. Restoring NF-E2 activity in a megakaryoblastic cell line or in NF-E2–deficient primary megakaryocytes rescues the expression of β1 tubulin. Re-expressing β1 tubulin in isolation does not, however, restore proplatelet formation in the defective megakaryocytes, indicating that other critical factors are required; indeed, other genes identified by mRNA subtraction also encode structural and regulatory components of the cytoskeleton. These findings provide critical mechanistic links between NF-E2, platelet formation, and selected microtubule proteins, and they also provide novel molecular insights into thrombopoiesis.
30
Boulabiar, Manel, Samir Boubaker, Michel Favre, and Caroline Demeret. "Keratinocyte sensitization to tumour necrosis factor-induced nuclear factor kappa B activation by the E2 regulatory protein of human papillomaviruses." Journal of General Virology 92, no. 10 (October 1, 2011): 2422–27. http://dx.doi.org/10.1099/vir.0.032466-0.
Full textAbstract:
Human papillomavirus (HPV) life cycle requires extensive manipulation of cell signalling to provide conditions adequate for viral replication within the stratified epithelia. In this regard, we show that the E2 regulatory protein of α, β and μ-HPV genotypes enhances tumour necrosis factor (TNF)-induced activation of nuclear factor kappa B (NF-κB). This activation is mediated by the N-terminal domain of E2, but does not rely on its transcriptional properties. It is independent of the NF-κB regulator Tax1BP1, which nevertheless interacts with all the E2 proteins. E2 specifically activates NF-κB pathways induced by TNF, while interleukin-1-induced pathways are not affected. E2 stimulates the activating K63-linked ubiquitination of TRAF5, and interacts with both TRAF5 and TRAF6. Our data suggest that E2 potentiates TNF-induced NF-κB signalling mediated by TRAF5 activation through direct binding. Since NF-κB controls epithelial differentiation, this activity may be involved in the commitment of infected keratinocytes to proliferation arrest and differentiation, both required for the implementation of the productive viral cycle.
31
Itoh, K., K. Igarashi, N. Hayashi, M. Nishizawa, and M. Yamamoto. "Cloning and characterization of a novel erythroid cell-derived CNC family transcription factor heterodimerizing with the small Maf family proteins." Molecular and Cellular Biology 15, no. 8 (August 1995): 4184–93. http://dx.doi.org/10.1128/mcb.15.8.4184.
Full textAbstract:
The chicken beta-globin enhancer is critical for the tissue- and developmental stage-specific expression of the beta-globin genes. This enhancer contains two indispensable cis elements, one containing two GATA sites and the other containing an NF-E2 site. To identify the putative transcription factor acting through the NF-E2 motif in the chicken beta-globin enhancer, we screened chicken cDNA libraries with a mouse p45 NF-E2 cDNA probe and isolated cDNA clones which encode a protein of 582 amino acid residues. This protein contains a region that includes the basic region-leucine zipper domain which is well conserved among members of the CNC family proteins (Cap 'n' collar, p45 NF-E2, LCR-F1, Nrf1, and Nrf2). Hence, we named this protein ECH (erythroid cell-derived protein with CNC homology). ECH is expressed abundantly in cultured erythroid cells undergoing terminal differentiation, peripheral erythrocytes, and some nonhematopoietic tissues. Since most of the cDNA clones obtained from the chicken erythrocyte cDNA library encoded ECH, ECH is likely the predominant CNC family protein present in avian peripheral erythrocytes. Like p45 NF-E2, ECH can heterodimerize with any of the small Maf family proteins and bind the NF-E2 site as a heterodimer in vitro. In a transfection assay, ECH transactivates transcription depending on the presence of NF-E2 sites on the reporter gene plasmid. These results indicate that ECH is likely a key regulator of avian erythropoiesis.
32
Aumann, Konrad, Anna-Verena Frey, Annette M. May, Dieter Hauschke, Clemens Kreutz, Jan P. Marx, Jens Timmer, Martin Werner, and Heike L. Pahl. "Subcellular mislocalization of the transcription factor NF-E2 in erythroid cells discriminates prefibrotic primary myelofibrosis from essential thrombocythemia." Blood 122, no. 1 (July 4, 2013): 93–99. http://dx.doi.org/10.1182/blood-2012-11-463257.
Full textAbstract:
Key Points The transcription factor NF-E2 is mislocalized in patients with primary myelofibrosis. Immunohistochemical staining for NF-E2 distinguishes essential thrombocythemia from primary myelofibrosis.
33
Wang, Wei, Bjorn W. Hackanson, and Heike L. Pahl. "Epigenetic Down-Regulation Of AML2 Expression By EZH2 Contributes To NF-E2 Overexpression In Myeloproliferative Neoplasms." Blood 122, no. 21 (November 15, 2013): 1605. http://dx.doi.org/10.1182/blood.v122.21.1605.1605.
Full textAbstract:
Abstract The transcription factor NF-E2 is overexpressed in the vast majority of patients with Myeloproliferative Neoplasms (MPN). In Essential Thrombocythemia (ET) and Primary Myelofibrosis (PMF) NF-E2 levels are elevated independent of the presence or absence of the JAK2V617Fmutation. We have recently shown that NF-E2 overexpression in a murine model leads to an MPN phenotype followed by spontaneous transformation to acute leukemia in a subset of mice. We have demonstrated that increased NF-E2 transcription is mediated by the transcription factor and proto-oncogene AML-1, which is overexpressed in MPN patients, again irrespective of the JAK2V617Fstatus. AML-1 binds to a conserved enhancer region located 3500 bp upstream of the NF-E2 transcriptional start site. Since AML-1 responsive genes are often also inversely regulated by the tumor suppressor AML-2 (RUNX3), we investigated whether NF-E2 expression is affected by AML-2 as well. Using chomatin immunoprecipitation assays (ChIP) we show that AML-2 binds the NF-E2 enhancer in vivo at a site distinct from but in proximity to the three AML-1 sites in the -3500bp region. AML-2 binding strongly represses transcription off the NF-E2 enhancer in reporter gene assays. This repression is abrogated by site directed mutagenesis of the AML-2 recognition sequence. Likewise, lentivirally induced AML-2 expression drastically reduces the amount of NF-E2 protein in erythroid and myeloid cells. These data clearly demonstrate that NF-E2 expression is directly regulated by AML-2. AML-2 thus serves as a repressor on the hematopoietic NF-E2 gene, a function previously noted mainly in solid tumors. In primary cells from patients with polycythemia vera (PV) AML-2 mRNA expression is significantly reduced. Moreover, ChIP assays demonstrate that in primary PV cells, significantly less AML-2 is bound to the NF-E2 enhancer than in healthy controls. Decreased repression by AML-2 thus cooperates with increased AML-1 induced transcription to elevate NF-E2 levels in PV patients. It has been demonstrated that AML-2 expression can be regulated by two distinct epigenetic mechanisms. For one, DNA methylation of the AML-2 promoter has been reported to silence AML-2 expression in gastric, colorectal and bladder cancers. On the other hand, aberrant histone methylation in the promoter region can also silence AML-2 expression. Here we show that DNA methylation of the AML-2 promoter is unaltered in PV patients. Rather, PV patients display aberrant histone methylation in the AML-2 promoter. Compared to healthy controls, H3K27me3 is significantly increased and H3K4me3 is significantly decreased in primary PV cells. This results in an inactive chromatin conformation on the AML-2 promoter in PV patients. Moreover, we show here that the histone-lysine-methyl-transferase “enhancer of zeste homologue 2” (EZH2), which confers the K3K27me3 mark, binds to the AML-2 promoter and decreases AML-2 expression. PV patients demonstrate significantly increased levels of EZH2 binding to the AML-2 promoter Treatment of primary PV cells and MPN cell lines with 2'-deoxy-5-azacytidine (DAC, Decitabine) ex vivo reverses the altered histone methylation, restoring the pattern found in healthy controls and decreases EZH2 binding to the AML-2 promoter. At the same time, Decitabine treatment induces AML-2 protein expression, decreases EZH2 expression and reduces the elevated NF-E2 levels. Moreover, a post-PV AML patient receiving Decitabine, displayed normalization of AML-2 promoter histone methylation and EZH2 binding on day 8 and day 15 of treatment. Taken together our data demonstrate that epigenetic silencing of AML-2 contributes to the elevated NF-E2 expression observed in MPN patients. Treatment with Decitabine restores physiological histone modifications to the AML-2 locus and decreases NF-E2 levels by reactivating AML-2 expression. These data provide a molecular rationale for extending the clinical investigation of epigenetic modifiers such as Decitabine or Azacitidine, currently used in MDS and AML, to patients with MPNs. A phase I study using Azacitidine in high risk PMF patients is currently being planned by the MPD-RC. Disclosures: No relevant conflicts of interest to declare.
34
Sharma, Ram V., Milind V. Gurjar, and Ramesh C. Bhalla. "Selected Contribution: Estrogen receptor-α gene transfer inhibits proliferation and NF-κB activation in VSM cells from female rats." Journal of Applied Physiology 91, no. 5 (November 1, 2001): 2400–2406. http://dx.doi.org/10.1152/jappl.2001.91.5.2400.
Full textAbstract:
Epidemiological studies have demonstrated that hormone replacement therapy with estrogen (E2) or E2plus progesterone in postmenopausal women decreases the age-associated risk of cardiovascular disease by 30–50%. Treatment of vascular smooth muscle (VSM) cells with physiological concentrations of E2 has been shown to inhibit growth factor-stimulated cell proliferation. In this study, we tested the hypothesis that E2 inhibits the age-associated increase in VSM cell proliferation by inhibiting nuclear factor (NF)-κB pathway. We investigated the effects of E2 treatment and adenovirus-mediated estrogen receptor (ER)-α gene transfer on cell proliferation and NF-κB activation using VSM cells cultured from 3-mo-old and 24-mo-old Fischer 344 female rats. Our results demonstrate that VSM cell proliferation was significantly increased ( P < 0.05) in aged compared with young adult female rats. Treatment of VSM cells with physiological concentrations of E2 inhibited VSM cell proliferation, and this inhibition was significantly greater ( P < 0.05) in cells from aged female rats compared with young adults. The inhibitory effects of E2 on cell proliferation in aged female rats were significantly potentiated by overexpression of the human ER-α gene into VSM cells. Constitutive and interleukin (IL)-1β-stimulated NF-κB activation was significantly greater ( P < 0.05) in VSM cells from aged compared with young female rats. E2 treatment of VSM cells from aged female rats inhibited both constitutive and IL-1β-stimulated NF-κB activation. ER-α gene transfer into VSM cells from aged female rats further augmented the inhibitory effects of E2. In conclusion, our data demonstrate that constitutive and IL-1β-stimulated NF-κB activation is increased in VSM cells from aged female rats due to loss of E2 and this can be restored back to normal levels by ER-α gene transfer and E2 treatment. In addition, increased NF-κB signaling may be responsible for increased incidence of cardiovascular disease in postmenopausal females.
35
Kataoka, K., K. Igarashi, K. Itoh, K. T. Fujiwara, M. Noda, M. Yamamoto, and M. Nishizawa. "Small Maf proteins heterodimerize with Fos and may act as competitive repressors of the NF-E2 transcription factor." Molecular and Cellular Biology 15, no. 4 (April 1995): 2180–90. http://dx.doi.org/10.1128/mcb.15.4.2180.
Full textAbstract:
The maf oncogene encodes a bZip nuclear protein which recognizes sequences related to an AP-1 site either as a homodimer or as heterodimers with Fos and Jun. We describe here a novel maf-related gene, mafG, which shows extensive homology with two other maf-related genes, mafK and mafF. These three maf-related genes encode small basic-leucine zipper proteins lacking the trans-activator domain of v-Maf. Bacterially expressed small Maf proteins bind to DNA as homodimers with a sequence recognition profile that is virtually identical to that of v-Maf. As we have previously described, the three small Maf proteins also dimerize with the large subunit of NF-E2 (p45) to form an erythroid cell-specific transcription factor, NF-E2, which has distinct DNA-binding specificity. This study shows that the small Maf proteins can also dimerize among themselves and with Fos and a newly identified p45-related molecule (Ech) but not with v-Maf or Jun. Although the small Maf proteins preferentially recognize the consensus NF-E2 sequence as heterodimers with either NF-E2 p45, Ech, or Fos, these heterodimers seemed to be different in their transactivation potentials. Coexpression of Fos and small Mafs could not activate a promoter with tandem repeats of the NF-E2 site. These results raise the possibility that tissue-specific gene expression and differentiation of erythroid cells are regulated by competition among Fos, NF-E2 p45, and Ech for small Maf proteins and for binding sites.
36
Pratt, M. A. Christine, Tanya E. Bishop, Dawn White, Gordon Yasvinski, Michel Ménard, Min Ying Niu, and Robert Clarke. "Estrogen Withdrawal-Induced NF-κB Activity and Bcl-3 Expression in Breast Cancer Cells: Roles in Growth and Hormone Independence." Molecular and Cellular Biology 23, no. 19 (October 1, 2003): 6887–900. http://dx.doi.org/10.1128/mcb.23.19.6887-6900.2003.
Full textAbstract:
ABSTRACT About one-third of breast cancers express a functional estrogen (β-estradiol [E2]) receptor (ER) and are initially dependent on E2 for growth and survival but eventually progress to hormone independence. We show here that ER+, E2-independent MCF-7/LCC1 cells derived from E2-dependent MCF-7 cells contain elevated basal NF-κB activity and elevated expression of the transcriptional coactivator Bcl-3 compared with the parental MCF-7 line. LCC1 NF-κB activity consists primarily of p50 dimers, although low levels of a p65/p50 complex are also present. The ER− breast cancer cell lines harbor abundant levels of both NF-κB complexes. In contrast, nuclear extracts from MCF-7 cells contain a significantly lower level of p50 and p65 than do LCC1 cells. Estrogen withdrawal increases both NF-κB DNA binding activity and expression of Bcl-3 in MCF-7 and LCC1 cells in vitro and in vivo. Tumors derived from MCF-7 cells ectopically expressing Bcl-3 remain E2 dependent but display a markedly higher tumor establishment and growth rate compared to controls. Expression of a stable form of IκBα in LCC1 cells severely reduced nuclear expression of p65 and the p65/p50 DNA binding heterodimer. Whereas LCC1 tumors in nude mice were stable or grew, LCC1(IκBα) tumors regressed after E2 withdrawal. Thus, both p50/Bcl-3- and p65/p50-associated NF-κB activities are activated early in progression and serve differential roles in growth and hormone independence, respectively. We propose that E2 withdrawal may initiate selection for hormone independence in breast cancer cells by activation of NF-κB and Bcl-3, which could then supplant E2 by providing both survival and growth signals.
37
Hirano, Suzue, Daisuke Furutama, and Toshiaki Hanafusa. "Physiologically high concentrations of 17β-estradiol enhance NF-κB activity in human T cells." American Journal of Physiology-Regulatory, Integrative and Comparative Physiology 292, no. 4 (April 2007): R1465—R1471. http://dx.doi.org/10.1152/ajpregu.00778.2006.
Full textAbstract:
Estrogen has diverse effects on inflammation and immune responses. That pregnancy is associated with remission of some autoimmune diseases and exacerbation of others suggests that physiological fluctuation in estrogen levels could affect the immune responses in humans. However, the molecular basis for these phenomena is poorly understood. We hypothesized that fluctuations of estrogen levels modulate intracellular signaling for immune responses via estrogen receptors (ERs). In reporter assays, 17β-estradiol (E2) at a physiologically high concentration increased the activity of NF-κB in Jurkat cells stimulated by PMA/ionomycin or TNF-α. Overexpression and RNA interference experiments suggested that the effects were mediated through ERβ. Immunoprecipitation assay showed that both ERα and ERβ are directly associated with NF-κB in the cell nucleus. Using chromatin immunoprecipitation assay, we confirmed that ERα and ERβ associated with NF-κB and steroid hormone coactivators at the promoter region of NF-κB regulated gene. Considering that NF-κB regulates the expression of various genes essential for cell growth and death, estrogen could regulate the fate of T cells by affecting the activity of NF-κB. To determine whether E2 alters the fate of T cells, we investigated E2 actions on T cell apoptosis, a well-known NF-κB-mediated phenomenon. E2 increased apoptosis of Jurkat cells and decreased that of human peripheral blood T cells. Our results indicate that E2 at a physiologically high concentration modulates NF-κB signaling in human T cells via ERβ and affects T cell survival, suggesting that these actions may underlie the gender differences in autoimmune diseases.
38
Palma, JF, X. Gao, CH Lin, S. Wu, and WB Solomon. "Iron protoporphyrin IX (hemin) but not tin or zinc protoporphyrin IX can stimulate gene expression in K562 cells from enhancer elements containing binding sites for NF-E2." Blood 84, no. 4 (August 15, 1994): 1288–97. http://dx.doi.org/10.1182/blood.v84.4.1288.1288.
Full textAbstract:
Abstract Many genes whose transcription is erythroid-specific contain enhancer or promoter elements that bind the transcription factor NF-E2. Hemin induction increases the expression of globin genes in the human erythroleukemia cell line K562, and increases the expression of reporters gene regulated by an enhancer elements containing binding sites for NF-E2. The failure of metalloporphyrins other than hemin to stimulate the transient expression of a CAT reporter gene linked to an enhancer element containing a binding site for NF-E2 was correlated with their failure to induce benzidine-positive K562 cells and increase the steady-state level of gamma-globin mRNA. This study suggests that elevated levels of zinc protoporphyrin IX found in the anemia of chronic disease, iron deficiency, and lead poisoning may contribute to a decrease in globin gene expression by interfering with the transcriptional activity of enhancer elements containing binding sites for NF-E2.
39
Palma, JF, X. Gao, CH Lin, S. Wu, and WB Solomon. "Iron protoporphyrin IX (hemin) but not tin or zinc protoporphyrin IX can stimulate gene expression in K562 cells from enhancer elements containing binding sites for NF-E2." Blood 84, no. 4 (August 15, 1994): 1288–97. http://dx.doi.org/10.1182/blood.v84.4.1288.bloodjournal8441288.
Full textAbstract:
Many genes whose transcription is erythroid-specific contain enhancer or promoter elements that bind the transcription factor NF-E2. Hemin induction increases the expression of globin genes in the human erythroleukemia cell line K562, and increases the expression of reporters gene regulated by an enhancer elements containing binding sites for NF-E2. The failure of metalloporphyrins other than hemin to stimulate the transient expression of a CAT reporter gene linked to an enhancer element containing a binding site for NF-E2 was correlated with their failure to induce benzidine-positive K562 cells and increase the steady-state level of gamma-globin mRNA. This study suggests that elevated levels of zinc protoporphyrin IX found in the anemia of chronic disease, iron deficiency, and lead poisoning may contribute to a decrease in globin gene expression by interfering with the transcriptional activity of enhancer elements containing binding sites for NF-E2.
40
Goerttler, Philipp S., Edith Maerz, Cordula Steimle, Britta Will, Annette Schmitt-Graeff, and Heike L. Pahl. "The Transcription Factor Nf-E2 Is Overexpressed in Patients with Polycythemia Vera." Blood 104, no. 11 (November 16, 2004): 659. http://dx.doi.org/10.1182/blood.v104.11.659.659.
Full textAbstract:
Abstract Despite recent advances in characterizing molecular markers for the diagnosis of polcycythemia vera (PV), the aberrations leading to disease development remain unknown. We therefore used expression profiling to identify candidate genes involved in the pathophysiology of PV. RNA from purified granulocytes of 40 PV patients was analyzed by hybridizing individual samples to a pool of 50 healthy controls. Of the 7,496 genes represented in the cDNA array, 253 were upregulated more than 1.5 fold in PV compared to healthy controls (p< 0.01, FDR corrected). Promoters for 26 of the 253 genes overexpressed in PV are regulated by members of the Sp1 family of transcription factors. We have therefore hypothesized that altered activity of one or several Sp1-like transcription factors may contribute to the molecular etiology of PV. Here we report that one of the Sp1 target genes identified, the transcription factor NF-E2, is overexpressed in 37 of the 40 PV patients (92.5%) assayed by microarray. NF-E2 overexpression was confirmed by Northern Blot and quantitative RT-PCR analysis. Transcription factor overexpression varies from 2.3 to 40 fold, with a median increase of 7 fold in PV patients compared to healthy controls. The NF-E2 protein is readily detected in PV granulocytes by Western Blot whereas it is undetectable in healthy control cells. Immunohistochemistry revealed that in PV bone marrow, NF-E2 is overexpressed in megakaryocytes as well as erythroid and granulocytic precursors. Several published observations suggest that NF-E2 is an exceptionally promising candidate in the molecular etiology of PV. Firstly, the transcription factor is expressed in hematopoietic precursors as well as in erythroid, megakaryocytic and granulocytic cells, those lineages affected in PV. Secondly, Sayer et al. have shown that overexpression of NF-E2 in fetal liver cells leads to the development of Epo-independent erythroid colonies, analogous to the endogenous erythroid colonies (EECs) observed in PV patients. Furthermore, ectopic expression of NF-E2 results in the spontaneous emergence of morphologically mature erythroid cells in the absence of Epo and can reprogram monocytic cells towards erythroid and megakaryocytic differentiation. These data support the hypothesis that the concentration of an individual transcription factor can control lineage commitment. We thus propose that in PV patients elevated concentrations of NF-E2 alter the physiological transcription factor balance leading to an overproduction of erythroid and, in select patients, megakaryocytic cells/platelets. In this model the level of NF-E2 overexpression determines both the severity of erythrocytosis and the concurrent presence or absence of thrombocytosis.
41
Imbrogno, Alessandra, Prantik Samanta, and Andrea I. Schäfer. "Fate of steroid hormone micropollutant estradiol in a hybrid magnetic ion exchange resin-nanofiltration process." Environmental Chemistry 16, no. 8 (2019): 630. http://dx.doi.org/10.1071/en19126.
Full textAbstract:
Environmental contextContamination of surface water by micropollutants is a major environmental concern because of their high persistence and toxicity. Micropollutants are only partially removed in nanofiltration water treatment systems, encouraging the investigation of more complex systems involving partitioning with membrane materials, organic matter and ion exchange resins. This study elucidates the micropollutant partitioning mechanisms in this complex water treatment system. AbstractThe accumulation of micropollutants, such as steroid hormones, in magnetic ion exchange resin-nanofiltration (MIEX-NF) poses a risk to the environmental contamination of surface water where the treated water is discharged. In this study, the partitioning of the steroid hormone estradiol (E2) with humic acid (HA), MIEX and the membrane is investigated at different feed water conditions (e.g. pH and presence of calcium). The transport and adsorption of E2 in NF is not affected significantly by the E2-HA interaction. Indeed, E2 partitions with HA between 8% and 25% at different pH. This is attributed to the presence of calcium ions, which reduces the number of HA molecules available to interact with E2 molecules. The calcium interference is evident especially at pH>10, where calcite and HA precipitate to result in irreversible membrane fouling. In the hybrid MIEX-NF process, the E2-MIEX interaction occurs at all pH conditions. Approximately 40% of the E2 total mass partitions with MIEX. This is significantly higher than E2 accumulation in NF. Since the partitioning is at least partially reversible, this poses a risk for accidental E2 release into the process streams.
42
Kapralova, Katarina, Lucie Lanikova, Felipe Lorenzo, Jihyun Song, Monika Horvathova, Vladimir Divoky, and Josef T. Prchal. "RUNX1 and NF-E2 upregulation is not specific for MPNs, but is seen in polycythemic disorders with augmented HIF signaling." Blood 123, no. 3 (January 16, 2014): 391–94. http://dx.doi.org/10.1182/blood-2013-10-534222.
Full textAbstract:
Key Points Overexpression of RUNX1 and its target NF-E2 is not specific for PV but is also seen in polycythemias due to augmented hypoxia sensing. Elevated levels of RUNX1 and NF-E2 are not specific for primary polycythemias, as these are not present in PFCP.
43
Goldenson, Benjamin, Qiang Jeremy Wen, and John D. Crispino. "Aurora A Kinase Is Required For Hematopoiesis and Couples Polyploidization With Terminal Differentiation In Megakaryocytes Through Phosphorylation Of NF-E2." Blood 122, no. 21 (November 15, 2013): 219. http://dx.doi.org/10.1182/blood.v122.21.219.219.
Full textAbstract:
Abstract We have recently shown that small molecule inhibitors of Aurora A kinase (AURKA) including dimethyfasudil and MLN827 induce polyploidization and differentiation of normal and malignant megakaryocytes, as evidenced by increased DNA content and upregulation of the cell surface markers CD41 and CD42. Furthermore, our pre-clinical studies demonstrate that MLN8237 shows potent anti-leukemia activity and anti-myelofibrotic activity in MPNs by promoting polyploidization and terminal differentiation of abnormal megakaryocytes. To determine the mechanism by which AURKA inhibitors ameliorate the leukemic and myelofibrotic phenotypes, we have examined the functional requirement for Aurka in adult hematopoiesis. To circumvent the embryonic lethality of the germline knockout and study the requirement for AURKA in adult hematopoiesis, we induced deletion in Aurka floxed mice with Mx1-Cre. Complete loss of AURKA caused a rapid and profound defect in hematopoiesis, with the mice developing pancytopenia and marked hypocellularity of the bone marrow. Notably, we observe an increase in the proportion of CD41 and CD42-positive megakaryocytes in contrast to the deficiency of myeloid and lymphoid cells in the bone marrow. To determine whether the observed defects are cell autonomous, we transplanted Aurkafl/fl, Mx1-Cre bone marrow cells to lethally irradiated recipients, confirmed engraftment, and induced deletion with pIpC. Upon AURKA deletion, the transplanted mice develop an identical phenotype to the one observed in the conditional knockout mice, demonstrating a cell autonomous requirement for AURKA during hematopoiesis. Moreover, in competitive transplantation experiments, Aurka-/- cells are selectively lost and fail to contribute to hematopoiesis. Mechanistically, loss of AURKA led to a significant degree of apoptosis in hematopoietic cells, likely due to mitotic catastrophe resulting from impaired chromosome segregation. To bypass the requirement for AURKA in progenitor cells and examine AURKA function in the megakaryocyte lineage, we deleted AURKA in megakaryocytes ex vivo by infecting Aurka floxed bone marrow with MSCV-Cre. We found that deletion of AURKA resulted in increased CD41 and CD42 expression as well as increased ploidy of the megakaryocyte fraction. Together these results are consistent with a selective differentiation effect of AURKA deficiency on megakaryocytes. To investigate whether AURKA modulates differentiation of megakaryocytes through interactions with lineage specific transcription factors, we performed co-immunoprecipitation experiments between AURKA and megakaryocyte transcription factors containing consensus AURKA phosphorylation sites in primary megakaryocytes. Results confirmed a robust interaction between AURKA and p45 NF-E2. Additionally, we found by an in vitro kinase assay that AURKA phosphorylates p45 NF-E2 on the S170 residue, as the S170A mutant of NF-E2 cannot be phosphorylated by AURKA. To explore the functional importance of NF-E2 phosphorylation at S170, we created a phosphomimetic mutant, S170D, and assayed its ability to induce megakaryocyte differentiation in primary cultures. While overexpression of wild-type NF-E2 significantly increased the expression of the megakaryocyte differentiation markers CD41 and CD42, overexpression of the S170D mutant was markedly less effective in promoting megakaryocyte differentiation, indicating that the loss of phosphorylation promotes NF-E2 activity and that NF-E2 activity may be suppressed by AURKA phosphorylation. Finally, to determine if AURKA inhibitors promote megakaryocyte differentiation by allowing activation of NF-E2, we knocked-down down NF-E2 in megakaryocytic cell lines and subsequently treated with diMF or MLN8237. Strikingly, cells with NF-E2 knocked-down displayed significantly less CD41 and CD42 expression in response to AURKA inhibitor treatment compared to control knockdowns. This experiment demonstrates that NF-E2 is required for the differentiation effect caused by AURKA inhibition. Taken together, our data show that Aurora A kinase is required for adult hematopoiesis and that Aurora A regulates NF-E2 function during megakaryocyte differentiation. Our results also reveal that inhibition of AURKA kinase activity couples polyploidization and terminal differentiation of megakaryocytes. Disclosures: Crispino: Sanofi: Research Funding.
44
Steiner, Laurie A., Yelena Maksimova, Vincent Schulz, Clara Wong, Debasish Raha, Milind C. Mahajan, Sherman M. Weissman, and Patrick G. Gallagher. "Chromatin Architecture and Transcription Factor Binding Regulate Expression of Erythrocyte Membrane Protein Genes." Molecular and Cellular Biology 29, no. 20 (August 17, 2009): 5399–412. http://dx.doi.org/10.1128/mcb.00777-09.
Full textAbstract:
ABSTRACT Erythrocyte membrane protein genes serve as excellent models of complex gene locus structure and function, but their study has been complicated by both their large size and their complexity. To begin to understand the intricate interplay of transcription, dynamic chromatin architecture, transcription factor binding, and genomic organization in regulation of erythrocyte membrane protein genes, we performed chromatin immunoprecipitation (ChIP) coupled with microarray analysis and ChIP coupled with massively parallel DNA sequencing in both erythroid and nonerythroid cells. Unexpectedly, most regions of GATA-1 and NF-E2 binding were remote from gene promoters and transcriptional start sites, located primarily in introns. Cooccupancy with FOG-1, SCL, and MTA-2 was found at all regions of GATA-1 binding, with cooccupancy of SCL and MTA-2 also found at regions of NF-E2 binding. Cooccupancy of GATA-1 and NF-E2 was found frequently. A common signature of histone H3 trimethylation at lysine 4, GATA-1, NF-E2, FOG-1, SCL, and MTA-2 binding and consensus GATA-1-E-box binding motifs located 34 to 90 bp away from NF-E2 binding motifs was found frequently in erythroid cell-expressed genes. These results provide insights into our understanding of membrane protein gene regulation in erythropoiesis and the regulation of complex genetic loci in erythroid and nonerythroid cells and identify numerous candidate regions for mutations associated with membrane-linked hemolytic anemia.
45
Kapralova, Katarina, Lucie Lanikova, Felipe R. Lorenzo V, Monika Horvathova, Vladimir Divoky, and Josef T. Prchal. "RUNX1 and NF-E2 Transcriptional Upregulation Is Not Specific For Myeloproliferative Neoplasms But Is Seen In Those Polycythemic Disorders With Augmented HIFs Signaling." Blood 122, no. 21 (November 15, 2013): 2177. http://dx.doi.org/10.1182/blood.v122.21.2177.2177.
Full textAbstract:
Abstract RUNX1 and NF-E2 are transcription factors that regulate hematopoietic stem cell homeostasis. It has been reported that increased RUNX1 expression in the granulocytes is present in all three classical myeloproliferative neoplasms (MPN): polycythemia vera (PV), essential thrombocythemia and primary myelofibrosis (Wang et al, Blood 2010), and that elevated NF-E2 promotes erythropoietin (EPO)-independent erythroid maturation of hematopoietic stem cells in vitro (Bogeska et al, Stem Cells Transl Med 2013). A mouse model overexpressing the NF-E2 transgene in hematopoietic cells was reported to be a new model of myeloproliferative neoplasms (Kaufmann et al, J Exp Med 2012). Polycythemic states can be divided into primary polycythemias, characterized by intrinsically hyperproliferative erythroid progenitors that are hypersensitive to EPO, and secondary polycythemias, wherein erythroid progenitors respond normally to EPO but circulating EPO is elevated or inappropriately normal for the level of increased red cell mass. Some congenital disorders including those with mutations in the hypoxia sensing pathway may share features of both primary and secondary polycythemias. We considered the possibility that increased transcripts of RUNX1 and NF-E2 might be the feature of other primary polycythemic states as well. We report a study of 19 polycythemic patients with primary or secondary polycythemia with diverse etiologies including mutations in positive and negative regulators of hypoxia sensing pathway. RUNX1 and NF-E2 transcripts were quantitated in granulocytes and BFU-E colonies by qPCR. All primary polycythemic patients had erythroid progenitors hypersensitive to or independent to EPO; all secondary polycythemic subjects had normal erythroid progenitor response to EPO. RUNX1 and NF-E2 gene transcripts were increased in granulocytes and BFU-E colonies in all PV patients, two unrelated subjects with the VHLR200W homozygous mutation (Chuvash polycythemia), one polycythemic patient homozygous for the VHLP138L exon 2 mutation, and a patient with the HIF2αM535V gain-of-function mutation. We also found upregulated expression of RUNX1 and NF-E2 in granulocytes and BFU-Es from a polycythemic patient (with no detectable EPOR, JAK2V617F or JAK2 exon 12 mutations and low level of EPO < 1 mU/mL) who was heterozygous for a SNP in exon 3 (rs147341899) in the LNK gene. We examined transcripts of RUNX1 and NF-E2 genes in granulocytes from two Croatian polycythemic patients with a homozygous VHLH191D exon 3 mutation whose erythroid progenitors were not hypersensitive to EPO and whose RUNX1, but not NF-E2, transcript was increased. We found similar results in two compound heterozygotes for VHLT124A exon 2 and VHLL188V exon 3 mutations. These two polycythemic siblings had hypersensitive erythroid colonies, increased RUNX1 transcripts and decreased NF-E2 transcripts in granulocytes. RUNX1 and NF-E2 transcripts were normal in two subjects with primary polycythemia due to the EPOR gain-of-function EPORQ434Xmutation, and in five unrelated subjects with secondary polycythemia. We next examined granulocyte transcripts of HIF-regulated genes: TFRC, SLC2A1, VEGF, BNIP3 and HK1, and found them to be increased in all PV patients and all studied polycythemic patients with VHL, HIF2α or LNK mutations, but not in polycythemic EPORQ434Xpatients or five patients with secondary polycythemia. Increased transcripts of HIF regulated genes are compatible with the previously unappreciated Warburg effect in PV (see S. Sana's Abstract at this ASH meeting). We propose that increased expression of RUNX1 and NF-E2 is not specific for myeloproliferative neoplasms but also is not universal for primary polycythemic disorders. Therefore, increased expression of RUNX1 and NF-E2 do not seem to be underlying mechanism for MPNs development but rather represent factors associated with diverse primary polycythemia states with augmented HIF signaling. (Note: KK and LL contributed equally to this work.) This work was supported by 1P01CA108671-O1A2 (NCI) Myeloproliferative Disorders (MPD) Consortium (PI Ron Hoffman) project#1 (PI Prchal) and the Leukemia & Lymphoma Society. Work by KK, LL, MH and VD was in part supported by the European Structural Funds (project CZ.1.07/2.3.00/20.0164 and CZ.1.07/2.3.00/30.0041), grant LF_2013_010 and by Czech Science Foundation (Project-P301/12/1503). Disclosures: No relevant conflicts of interest to declare.
46
McCormack, Matthew P., Mark A. Hall, Simone M. Schoenwaelder, Quan Zhao, Sarah Ellis, Julia A. Prentice, Ashleigh J. Clarke, et al. "A critical role for the transcription factor Scl in platelet production during stress thrombopoiesis." Blood 108, no. 7 (October 1, 2006): 2248–56. http://dx.doi.org/10.1182/blood-2006-02-002188.
Full textAbstract:
Abstract The generation of platelets from megakaryocytes in the steady state is regulated by a variety of cytokines and transcription factors, including thrombopoietin (TPO), GATA-1, and NF-E2. Less is known about platelet production in the setting of stress thrombopoiesis, a pivotal event in the context of cytotoxic chemotherapy. Here we show in mice that the transcription factor Scl is critical for platelet production after chemotherapy and in thrombopoiesis induced by administration of TPO. Megakaryocytes from these mice showed appropriate increases in number and ploidy but failed to shed platelets. Ultrastructural examination of Scl-null megakaryocytes revealed a disorganized demarcation membrane and reduction in platelet granules. Quantitative real-time polymerase chain reaction showed that Scl-null platelets lacked NF-E2, and chromatin immunoprecipitation analysis demonstrated Scl binding to the NF-E2 promoter in the human megakaryoblastic-cell line Meg-01, along with its binding partners E47, Lmo2, and the cofactors Ldb1 and GATA-2. These findings suggest that Scl acts up-stream of NF-E2 expression to control megakaryocyte development and platelet release in settings of thrombopoietic stress.
47
Ackley, Catherine J., and Christopher H. Lowrey. "GATA-1, NF-E2 and EKLF/SP1 Binding Elements Cooperate To Specify Patterns of Histone Hyper-Acetylation at β-Globin LCR HS Core Elements." Blood 104, no. 11 (November 16, 2004): 1208. http://dx.doi.org/10.1182/blood.v104.11.1208.1208.
Full textAbstract:
Abstract The active elements of the β-globin LCR are necessary for high-level expression of the linked globin genes. These active elements are contained within regions of unique, erythroid-specific chromatin structure characterized by the formation of positioned nucleosomal arrays, areas of approximately 150–200 bp which are highly accessible for protein-DNA interactions (DNase I HSs) and hyperacetylation of histones H3 and H4. At a DNA level, each of the LCR HS cores contain NF-E2, EKLF/Sp1 and tandem, inverted GATA binding elements. Our lab has shown that all of these transcription factor-binding sites are necessary for HS formation. We have previously proposed a model of HS formation in which Sp1 and/or EKLF first bind to the core regions and alter local chromatin structure allowing GATA-1 and NF-E2 to bind and stabilize the HS core structure. Additional factors could then bind to the core and flanking regions. An important aspect of HS core formation that our experiments have not previously addressed is the mechanism of histone acetylation. Based on our previous findings and the fact that GATA-1 and NF-E2 can both interact with histone acetyltransferases (HATs), we hypothesized that histone acetylation at the HS requires the binding of EKLF/Sp1 followed by GATA-1 and/or NF-E2 and therefor should be a late event in HS formation. To test this hypothesis, we have generated a series of 11 artificial HS core elements based on the structure of human HS4. These plasmid-based constructs contain different combinations of the six binding sites in their normal arrangement and spacing: (EKLF/Sp1)-(NF-E2)-(EKLF/SP1)-(GATA-1 x 2)-(EKLF/Sp1). These constructs were stably transfected into MEL cells and assayed for their ability to mediate histone H3 and H4 acetylation as determined by the ChIP assay. The endogenous murine β-globin LCR HS3 served as a positive internal control for histone acetylation. Previous results from our lab showed that all six factor-binding sites within the HS core are required for HS formation. However, when assayed for histone acetylation, the NF-E2 and the tandem inverted GATA sites are able to independently direct H3 and H4 acetylation to between 10 and 50% of that seen at the native HS site. Combination of the NF-E2 and GATA-1 sites results in 89% of normal H4 acetylation. Full HH4 acetylation (98% of the wild-type HS) is only seen with all six factor binding sites. These results indicate that NF-E2 and GATA-1 direct most of the acetylation of LCR HS core histones but that EKLF binding is also likely to contribute. They also indicate that extensive histone acetylation can occur without HS formation. Finally, in contrast to our model of HS formation, GATA-1 and NF-E2 appear able to direct the acetylation of HS core histones without the presence of binding sites for Sp1 or EKLF.
48
Ritter, Clare L., William F. Prigge, Mark A. Reichert, and Danuta Malejka-Giganti. "Oxidations of 17β-estradiol and estrone and their interconversions catalyzed by liver, mammary gland and mammary tumor after acute and chronic treatment of rats with indole-3-carbinol or β-naphthoflavone." Canadian Journal of Physiology and Pharmacology 79, no. 6 (June 1, 2001): 519–32. http://dx.doi.org/10.1139/y01-020.
Full textAbstract:
Altered cytochrome P450-catalyzed metabolism of 17β-estradiol (E2) and estrone (E1) in the liver and (or) extrahepatic tissues may affect estrogen-sensitive tumorigenesis. We examined the effects of oral treatments of (i) indole-3-carbinol (I3C) at 250 or 500 mg/kg or β-naphthoflavone (β-NF) at 40 mg/kg of body weight (bw)/day from 51 to 54 days of age (acute regimen), and (ii) I3C at 250 mg/kg or β-NF at 20 mg/kg bw given 3x/week from 10 to 22 weeks of age (chronic regimen) in female Sprague-Dawley rats. We determined the effects of these treatments on the P450 content and P450 (CYP)-specific activities in the liver, P450-dependent metabolism of E2 and E1 by the liver and mammary gland, and interconversion of E1 and E2 catalyzed by 17β-hydroxysteroid dehydrogenase (17β-HSD) in these tissues and malignant mammary tumors. I3C at the two levels of acute regimen elicited similar responses. Acute and chronic treatments with I3C, but not β-NF, increased P450 content ~2-fold. I3C, and to a lesser extent β-NF, increased CYP1A1 and CYP1A2 probe activities in liver up to 117- and 27- fold, respectively, and after acute regimens, that of CYP3A by ~1.8-fold. I3C also increased activity of CYP2B up to 100-fold. Overall hepatic metabolism of E2 and E1, which was ~2-fold greater at 55 than 155 days of age, was increased (~2.8-fold) by I3C with 2-, 4-, 16α-, 6α-, 6β-, and 15α-hydroxy (OH) comprising [Formula: see text]54, 3, 2, ~2, ~5, 7, and 2%, respectively, of E1 and E2 metabolites. Acute regimens of β-NF increased 2- and 15α-OH-E2 (62 and 5% of total) from E2 and 2-, 4-, and 6α-OH-E1 + 6β-OH-E1 (32, 13, and 4% of total) from E1. Mammary gland metabolized E2 to E1 and small amounts of 15α-, 4-, 16α-, 6β-, and 6α-OH-E2. After the acute IC3 regimen, E2 was also converted to 2-OH-E2. 17β-HSD-catalyzed oxidation of E2 was favored in the liver and reduction of E1 was favored in mammary gland and tumor (= 1% of hepatic activity). An increased (~2-fold) ratio of reductive to oxidative activities in malignant mammary tumors by chronic I3C regimen may stimulate tumor growth. This is the first report showing that after chronic oral regimens, the I3C-, but not β-NF-, induced changes in CYP complement led to elevated E2 and E1 metabolism. The persistent effects of increased putative carcinogenic and estrogenic 4- and 16α-OH as well as 6α- and 6β-OH-E2 and 6β-OH-E1 might counteract those of the less estrogenic 2-OH metabolites, thus accounting for the lack of suppression of mammary tumorigenesis by I3C in our previous study.Key words: estrogen metabolism, P450, 17β-hydroxysteroid dehydrogenase, indole-3-carbinol, β-naphthoflavone.
49
Andrews, Nancy C. "Erythroid Transcription Factor NF-E2 Coordinates Hemoglobin Synthesis." Pediatric Research 36, no. 4 (October 1994): 419–23. http://dx.doi.org/10.1203/00006450-199410000-00001.
Full text
50
Andrews, Nancy C. "Molecules in focus The NF-E2 transcription factor." International Journal of Biochemistry & Cell Biology 30, no. 4 (April 1998): 429–32. http://dx.doi.org/10.1016/s1357-2725(97)00135-0.
Full text