scholarly journals Core binding factor cannot synergistically activate the myeloperoxidase proximal enhancer in immature myeloid cells without c-Myb.

1997 ◽  
Vol 17 (9) ◽  
pp. 5127-5135 ◽  
Author(s):  
M Britos-Bray ◽  
A D Friedman
Keyword(s):  
Binding Site ◽  
Binding Sites ◽  
Myeloid Cells ◽  
Core Binding Factor ◽  
Enhancer Activity ◽  
Binding Factor ◽  
Evolutionarily Conserved ◽  
Immature Myeloid Cells ◽  
Stimulating Factor ◽  
Deletion Series

The myeloperoxidase (MPO) gene is transcribed specifically in immature myeloid cells and is regulated in part by a 414-bp proximal enhancer. Mutation of a core binding factor (CBF)-binding site at -288 decreased enhancer activity 30-fold in 32D cl3 myeloid cells cultured in granulocyte colony-stimulating factor (G-CSF). A novel functional analysis, linking the CBF-binding site to an enhancer deletion series, located at -147 an evolutionarily conserved c-Myb-binding site which was required for optimal enhancer activity and synergy with CBF in 32D cells. These sites cooperated in isolation and independent of a precise spacing. Deletional analysis carried out in the absence of the c-Myb-binding site at -147 located at -301 a second c-Myb-binding site which also synergized with CBF to activate the enhancer. A GA-rich region at -162 contributed to cooperation with CBF when the adjacent c-Myb-binding site was intact. Mutation of both c-Myb-binding sites in the context of the entire enhancer greatly impaired activation by endogenous CBF in 32D cells. Similarly, activation by c-Myb was impaired in constructs lacking the CBF-binding site. CBF and c-Myb were required for induction of MPO proximal enhancer activity when 32D cells differentiated in response to G-CSF. A fusion protein containing the Gal4 DNA-binding domain and the AML-1B activation domain, amino acids 216 to 480, activated transcription alone and cooperatively with c-Myb in nonmyeloid CV-1 cells. Determining how CBF and c-Myb synergize in myeloid cells might contribute to our understanding of leukemogenesis by the AML1-ETO, AML1-MDS1, CBFbeta-SMMHC, and v-Myb oncoproteins.

scholarly journals Interleukin 1α (IL-1α) Promotes Pathogenic Immature Myeloid Cells and IL-1β Favors Protective Mature Myeloid Cells During Acute Lung Infection

2018 ◽  
Vol 217 (9) ◽  
pp. 1481-1490 ◽  
Author(s):  
Sivakumar Periasamy ◽  
Jonathan A Harton
Keyword(s):  
Interleukin 1 ◽  
Myeloid Cells ◽  
Myeloid Cell ◽  
Interleukin 10 ◽  
Dual Function ◽  
Interleukin 17 ◽  
Colony Stimulating Factor ◽  
Immature Myeloid Cells ◽  
Severe Pathology ◽  
Stimulating Factor

Abstract Bacterial pneumonia is a common risk factor for acute lung injury and sepsis-mediated death, but the mechanisms underlying the overt inflammation and accompanying pathology are unclear. Infiltration of immature myeloid cells and necrotizing inflammation mediate severe pathology and death during pulmonary infection with Francisella tularensis. However, eliciting mature myeloid cells provides protection. Yet, the host factors responsible for this pathologic immature myeloid cell response are unknown. Here, we report that while the influx of both mature and immature myeloid cells is strictly MyD88 dependent, the interleukin 1 (IL-1) receptor mediates an important dual function via its ligands IL-1α and IL-1β. Although IL-1β favors the appearance of bacteria-clearing mature myeloid cells, IL-1α contributes to lung infiltration by ineffective and pathologic immature myeloid cells. Finally, IL-1α and IL-1β are not the sole factors involved, but myeloid cell responses during acute pneumonia were largely unaffected by lung levels of interleukin 10, interleukin 17, CXCL1, granulocyte colony-stimulating factor, and granulocyte-macrophage colony-stimulating factor.

C-MYB and C/EBP Family Transcription Factors Regulate the Gfi-1 Promoter.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 4214-4214
Author(s):  
Richard Dahl ◽  
Kristin S. Owens
Keyword(s):  
Transcription Factor ◽  
Transcription Factors ◽  
Chromatin Immunoprecipitation ◽  
Binding Sites ◽  
Myeloid Cells ◽  
Transient Transfection ◽  
Mobility Shift ◽  
Reporter Assays ◽  
Immature Myeloid Cells ◽  
Factor C

Abstract Gfi-1 −/− mice generate abnormal immature myeloid cells exhibiting characteristics of both monocytes and granulocytes. One of Gfi-1’s critical functions is to downregulate monocyte specific genes in order for granulocytes to develop properly. Since the transcription factors C/EBP alpha and C/EBP epsilon are needed for granulocyte development we hypothesized that these factors may regulate Gfi-1 expression. The Gfi-1 promoter contains several putative C/EBP binding sites and we show by electrophoretic mobility shift and chromatin immunoprecipitation that C/EBP family members can bind to some of these sites. However we were unable to see activation of the Gfi-1 promoter by C/EBP proteins in transient transfection reporter assays. Other groups have shown that C/EBP proteins can synergize with the transcription factor c-myb. We observed that the Gfi-1 promoter contains sites for the hematopoietic transcription factor c-myb. Sevral of these c-myb binding sites are adjacent to C/EBP binding sites. In reporter assays in non-hematopoietic cells c-myb activated the Gfi-1 promoter by itself and this activity was enhanced when we included either C/EBP alpha or epsilon in the transfection. Our data suggests that C/EBP proteins and c-myb regulate the transcription of Gfi-1 in myeloid cells.

The prothrombin 3′end formation signal reveals a unique architecture that is sensitive to thrombophilic gain-of-function mutations

Blood ◽  
2004 ◽  
Vol 104 (2) ◽  
pp. 428-435 ◽  
Author(s):  
Sven Danckwardt ◽  
Niels H. Gehring ◽  
Gabriele Neu-Yilik ◽  
Patrick Hundsdoerfer ◽  
Margit Pforsich ◽  
...  
Keyword(s):  
Binding Site ◽  
Binding Sites ◽  
Flanking Sequence ◽  
Gain Of Function ◽  
Functional Efficiency ◽  
Processing Signal ◽  
The Common ◽  
Low Efficiency ◽  
Stimulating Factor

Abstract The functional analysis of the common prothrombin 20210 G>A(F2 20210*A) mutation has recently revealed gain of function of 3′end processing as a novel genetic mechanism predisposing to human disease. We now show that the physiologic G at the cleavage site at position 20210 is the functionally least efficient nucleotide to support 3′end processing but has evolved to be physiologically optimal. Furthermore, the F2 3′end processing signal is characterized by a weak downstream cleavage stimulating factor (CstF) binding site with a low uridine density, and the functional efficiency of F2 3′end processing can be enhanced by the introduction of additional uridine residues. The recently identified thrombosis-related mutation (F2 20221*T) within the CstF binding site up-regulates F2 3′end processing and prothrombin biosynthesis in vivo. F2 20221*T thus represents the first example of a likely pathologically relevant mutation of the putative CstF binding site in the 3′flanking sequence of a human gene. Finally, we show that the low-efficiency F2 cleavage and CstF binding sites are balanced by a stimulatory upstream uridine-rich element in the 3′UTR. The architecture of the F2 3′end processing signal is thus characterized by a delicate balance of positive and negative signals. This balance appears to be highly susceptible to being disturbed by clinically relevant gain-of-function mutations. (Blood. 2004;104:428-435)

scholarly journals C/EBP, c-Myb, and PU.1 cooperate to regulate the neutrophil elastase promoter.

1996 ◽  
Vol 16 (9) ◽  
pp. 4717-4725 ◽  
Author(s):  
M Oelgeschläger ◽  
I Nuchprayoon ◽  
B Lüscher ◽  
A D Friedman
Keyword(s):  
Neutrophil Elastase ◽  
Binding Sites ◽  
Myeloid Cells ◽  
3T3 Cells ◽  
Transactivation Domains ◽  
Dna Binding Sites ◽  
Immature Myeloid Cells ◽  
Nih 3T3 ◽  
Cooperative Activation ◽  
Nih 3T3 Cells

The murine neutrophil elastase (NE) gene is expressed specifically in immature myeloid cells. A 91-bp NE promoter region contains three cis elements which are conserved evolutionarily and are essential for activation of the promoter in differentiating 32D cl3 myeloid cells. These elements bound c-Myb (at -49), C/EBPalpha (at -57), and PU.1 (at -82). In NIH 3T3 cells, the NE promoter was activated by c-Myb, C/EBPalpha, and PU.1, via their respective binding sites. Cooperative activation was seen by any combination of c-Myb, C/EBPalpha, and PU.1, including all three together, again via their DNA-binding sites. In CV-1 cells, but not in NIH 3T3 cells, cooperation between Myb and C/EBPalpha depended on the integrity of the PU.1-binding site. In addition to C/EBPalpha, C/EBPdelta strongly activated the NE promoter, alone or with c-Myb, but C/EBPbeta was less active. Either of C/EBPalpha's two transactivation domains cooperatively activated the promoter with c-Myb, in both NIH 3T3 and 32D c13 cells. Synergistic binding to DNA in a gel shift assay between C/EBPalpha, c-Myb, and PU.1 could not be demonstrated. Also, separation of the C/EBP- and c-Myb-binding sites by 5 or 10 bp did not prevent cooperativity. These results suggest that a coactivator protein mediates cooperative activation of the NE promoter by a C/EBP and c-Myb. These factors, together with PU.1, direct restricted expression of the NE promoter to immature myeloid cells.

Hepatocyte nuclear factor 1 and C/EBP are essential for the activity of the human apolipoprotein B gene second-intron enhancer

1992 ◽  
Vol 12 (3) ◽  
pp. 1134-1148
Author(s):  
A R Brooks ◽  
B Levy-Wilson
Keyword(s):  
Binding Site ◽  
Binding Sites ◽  
Apolipoprotein B ◽  
Nuclear Factor ◽  
Enhancer Activity ◽  
Protein Binding Sites ◽  
Human Apolipoprotein ◽  
Nuclear Factor 1 ◽  
Related Proteins ◽  
Apolipoprotein B Gene

The tissue-specific transcriptional enhancer of the human apolipoprotein B gene contains multiple protein-binding sites spanning 718 bp. Most of the enhancer activity is found in a 443-bp fragment (+621 to +1064) that is located entirely within the second intron of the gene. Within this fragment, a 147-bp region (+806 to +952) containing a single 97-bp DNase I footprint exhibits significant enhancer activity. We now report that this footprint contains four distinct protein-binding sites that have the potential to bind nine distinct liver nuclear proteins. One of these proteins was identified as hepatocyte nuclear factor 1 (HNF-1), which binds with relatively low affinity to the 5' half of a 20-bp palindrome located at the 5' end of the large footprint. A binding site for C/EBP (or one of the related proteins that recognize similar sequences) was identified in the center of the 97-bp footprint. This binding site is coincident or overlaps with the binding sites for five other proteins, two of which appear to be distinct from the C/EBP-related family of proteins. The binding site for a nuclear factor designated protein I is located between the HNF-1 and C/EBP binding sites. Finally, the 3'-most 15 bp of the footprinted sequence contain a binding site for another nuclear protein, which we have called protein II. Mutations that abolish the binding of either HNF-1, protein II, or the C/EBP-related proteins severely reduce enhancer activity. However, deletion experiments demonstrated that neither the HNF-1-binding site alone, nor the combination of binding sites for HNF-1, protein I, and C/EBP, nor the C/EBP-binding site plus the protein II-binding site is sufficient to enhance transcription from a strong apolipoprotein B promoter. Rather, HNF-1 and C/EBP act synergistically with protein II to enhance transcription of the apolipoprotein B gene.

scholarly journals Identification and characterization of Hoxa9 binding sites in hematopoietic cells

Blood ◽  
2012 ◽  
Vol 119 (2) ◽  
pp. 388-398 ◽  
Author(s):  
Yongsheng Huang ◽  
Kajal Sitwala ◽  
Joel Bronstein ◽  
Daniel Sanders ◽  
Monisha Dandekar ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Binding Sites ◽  
Enhancer Activity ◽  
Transcription Start Sites ◽  
Genome Wide ◽  
A Genome ◽  
Evolutionarily Conserved ◽  
P300 Binding ◽  
Histone H3k4

The clustered homeobox proteins play crucial roles in development, hematopoiesis, and leukemia, yet the targets they regulate and their mechanisms of action are poorly understood. Here, we identified the binding sites for Hoxa9 and the Hox cofactor Meis1 on a genome-wide level and profiled their associated epigenetic modifications and transcriptional targets. Hoxa9 and the Hox cofactor Meis1 cobind at hundreds of highly evolutionarily conserved sites, most of which are distant from transcription start sites. These sites show high levels of histone H3K4 monomethylation and CBP/P300 binding characteristic of enhancers. Furthermore, a subset of these sites shows enhancer activity in transient transfection assays. Many Hoxa9 and Meis1 binding sites are also bound by PU.1 and other lineage-restricted transcription factors previously implicated in establishment of myeloid enhancers. Conditional Hoxa9 activation is associated with CBP/P300 recruitment, histone acetylation, and transcriptional activation of a network of proto-oncogenes, including Erg, Flt3, Lmo2, Myb, and Sox4. Collectively, this work suggests that Hoxa9 regulates transcription by interacting with enhancers of genes important for hematopoiesis and leukemia.

scholarly journals Hepatocyte nuclear factor 1 and C/EBP are essential for the activity of the human apolipoprotein B gene second-intron enhancer.

1992 ◽  
Vol 12 (3) ◽  
pp. 1134-1148 ◽  
Author(s):  
A R Brooks ◽  
B Levy-Wilson
Keyword(s):  
Binding Site ◽  
Binding Sites ◽  
Apolipoprotein B ◽  
Nuclear Factor ◽  
Enhancer Activity ◽  
Protein Binding Sites ◽  
Human Apolipoprotein ◽  
Nuclear Factor 1 ◽  
Related Proteins ◽  
Apolipoprotein B Gene

The tissue-specific transcriptional enhancer of the human apolipoprotein B gene contains multiple protein-binding sites spanning 718 bp. Most of the enhancer activity is found in a 443-bp fragment (+621 to +1064) that is located entirely within the second intron of the gene. Within this fragment, a 147-bp region (+806 to +952) containing a single 97-bp DNase I footprint exhibits significant enhancer activity. We now report that this footprint contains four distinct protein-binding sites that have the potential to bind nine distinct liver nuclear proteins. One of these proteins was identified as hepatocyte nuclear factor 1 (HNF-1), which binds with relatively low affinity to the 5' half of a 20-bp palindrome located at the 5' end of the large footprint. A binding site for C/EBP (or one of the related proteins that recognize similar sequences) was identified in the center of the 97-bp footprint. This binding site is coincident or overlaps with the binding sites for five other proteins, two of which appear to be distinct from the C/EBP-related family of proteins. The binding site for a nuclear factor designated protein I is located between the HNF-1 and C/EBP binding sites. Finally, the 3'-most 15 bp of the footprinted sequence contain a binding site for another nuclear protein, which we have called protein II. Mutations that abolish the binding of either HNF-1, protein II, or the C/EBP-related proteins severely reduce enhancer activity. However, deletion experiments demonstrated that neither the HNF-1-binding site alone, nor the combination of binding sites for HNF-1, protein I, and C/EBP, nor the C/EBP-binding site plus the protein II-binding site is sufficient to enhance transcription from a strong apolipoprotein B promoter. Rather, HNF-1 and C/EBP act synergistically with protein II to enhance transcription of the apolipoprotein B gene.

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