C/EBPalpha Is a Key Transcriptional Regulator of the Triggering Receptor on Myeloid Cells-2 Gene

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 4762-4762
Author(s):  
Maria Michela Mancarelli ◽  
Lily Catherine Frusciante ◽  
Gabrielle Francoise Cauvi ◽  
Charles De Rossi ◽  
Bruce Edward Torbett
Keyword(s):  
Gene Regulation ◽  
Binding Site ◽  
Binding Sites ◽  
Transcriptional Control ◽  
Transcriptional Activity ◽  
Myeloid Cells ◽  
Myeloid Cell ◽  
Fold Increase ◽  
Full Length ◽  
And Function

Abstract Triggering receptor on myeloid cells-2 (TREM-2) is a member of the innate immune signaling receptor TREM family. After differentiation of monocytes, TREM-2 is expressed on the cell surface of macrophages, monocyte-derived dendritic cells, microglia, and osteoclasts. TREM-2 is necessary for modulation of cellular activation and function, and loss of TREM-2 results in the decrease of phagocytosis of apoptotic neurons and cells and an increase in inflammation response of macrophage-lineage cells. To better understand TREM-2 gene regulation during monocyte differentiation we isolated and then analyzed the putative human TREM-2 promoter to identify transcriptional control. The 1131 bp TREM- 2 promoter contains binding sites for C/EBPα and PU.1 transcription factors, known for regulation of myeloid cell differentiation and function. A series of 5 promoter reporter deletion mutants were generated to dissect transcription regulation. We progressively deleted the full-length promoter, −1002/+129, of HIF1, STAT5, GATA1, YY1F, C/EBPα binding sites generating −866/+129, −632/+129, −526/+129, −258/+129, +37/+129 promoter reporters, respectively. All TREM-2 promoter reporters showed minimal transcriptional activity when transfected in 293T-HEK cells. In contrast, C/EBPα was necessary and sufficient for TREM-2 promoter reporter activity in co-transfection studies, demonstrating a 5-fold induction of transcriptional activity as compared to controls. Consistent with the TREM-2 reporter findings showing C/EBPα mediated activity, EMSA assays demonstrated specific C/EBPα binding to sites within the TREM-2 promoter. To determine if both C/EBPα binding sites in TREM-2 were necessary for activity we generated loss of C/EBPα binding site reporters and observed that only the −43/−28 C/EBPα binding site was essential for activity. To clarify if +73/+93 PU.1 binding site had a role in TREM-2 regulation, we co-transfected the full-length promoter with PU.1 and C/EBPα and observed a 10-fold increase of induction of TREM-2 transcriptional activity. However, PU.1 alone did not activate the full-length TREM-2 promoter or bind to the PU.1 consensus site, and loss of the PU.1 binding site did not alter transcriptional activity. These results suggest a key role for direct C/EBPα and indirect PU.1 participation in regulation of TREM-2. Studies are underway to precisely understand TREM-2 gene regulation and how TREM-2 controls macrophage-mediated phagocytosis and inflammation.

The rat albumin promoter: cooperation with upstream elements is required when binding of APF/HNF1 to the proximal element is partially impaired by mutation or bacterial methylation

1989 ◽  
Vol 9 (11) ◽  
pp. 4759-4766
Author(s):  
F Tronche ◽  
A Rollier ◽  
I Bach ◽  
M C Weiss ◽  
M Yaniv
Keyword(s):  
Binding Site ◽  
Binding Sites ◽  
Transcriptional Activity ◽  
Transient Transfection ◽  
Tata Box ◽  
Specific Factor ◽  
Target Sequence ◽  
Functional Interaction ◽  
Dna Methylase ◽  
Ccaat Box

We have characterized in the accompanying paper (P. Herbomel, A. Rollier, F. Tronche, M.-O. Ott, M. Yaniv, and M. C. Weiss, Mol. Cell. Biol. 9:4750-4758, 1989) six different elements in the albumin promoter. One of them, the proximal element (PE), is the binding site for a strictly liver specific factor, APF/HNF1. This binding site contains a bacterial DAM DNA methylase methylation target sequence which, when methylated, decreases the affinity of the protein for this element. When the different albumin promoter constructions were prepared in an Escherichia coli deoxyadenosine methylase-negative strain, the respective contributions of the elements to the overall promoter activity were strikingly different. An intact proximal element plus the TATA box gave almost full transcriptional activity in transient transfection experiments and only in differentiated hepatoma cells of line H4II, whereas the distal elements (distal element III [DEIII], the NF1-binding site DEII, and the E/CBP-binding site DEI) had become essentially dispensable. Mutations affecting the CCAAT box showed only a two- to threefold decrease. When PE was methylated, mutated, or replaced by the homologous element from the alpha-fetoprotein gene, activity in the context of the short promoter (PE plus the TATA box) was abolished. However, activity was restored in the presence of the upstream elements, showing that cooperation with factors binding to the CCAAT box and distal elements favors the functional interaction of the liver-specific APF/HNF1 factor with lower-affinity binding sites.

scholarly journals Regulation of rice root development by a retrotransposon acting as a microRNA sponge

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Jungnam Cho ◽  
Jerzy Paszkowski
Keyword(s):  
Gene Regulation ◽  
Transcription Factors ◽  
Root Development ◽  
Binding Sites ◽  
Transcriptional Control ◽  
Specific Expression ◽  
Ability To Act ◽  
Specific Accumulation ◽  
In Trans ◽  
Microrna Sponge

It is well documented that transposable elements (TEs) can regulate the expression of neighbouring genes. However, their ability to act in trans and influence ectopic loci has been reported rarely. We searched in rice transcriptomes for tissue-specific expression of TEs and found them to be regulated developmentally. They often shared sequence homology with co-expressed genes and contained potential microRNA-binding sites, which suggested possible contributions to gene regulation. In fact, we have identified a retrotransposon that is highly transcribed in roots and whose spliced transcript constitutes a target mimic for miR171. miR171 destabilizes mRNAs encoding the root-specific family of SCARECROW-Like transcription factors. We demonstrate that retrotransposon-derived transcripts act as decoys for miR171, triggering its degradation and thus results in the root-specific accumulation of SCARECROW-Like mRNAs. Such transposon-mediated post-transcriptional control of miR171 levels is conserved in diverse rice species.

Overexpression of CCAAT Displacement Protein Represses the Promiscuously Active Proximal gp91phox Promoter

Blood ◽  
1999 ◽  
Vol 94 (9) ◽  
pp. 3151-3160 ◽  
Author(s):  
Diana Catt ◽  
Shannon Hawkins ◽  
Ann Roman ◽  
Wen Luo ◽  
David G. Skalnik
Keyword(s):  
Binding Site ◽  
Binding Sites ◽  
Transcriptional Activity ◽  
Binding Activity ◽  
Proximal Promoter ◽  
Modular Organization ◽  
Transcriptional Activators ◽  
Cis Element ◽  
Dna Binding Activity ◽  
Ccaat Displacement Protein

CCAAT displacement protein (CDP) is a transcriptional repressor that restricts expression of the gp91phox gene to mature myeloid cells. CDP interacts with multiple sites within the −450 to +12 bp human gp91phox promoter, and down-regulation of CDP DNA-binding activity is required for induction of gp91phox transcription in mature phagocytes. Truncation of the gp91phox promoter to −102 to +12 bp removes 4 CDP-binding sites and reveals a promiscuous promoter activity that is active in some nonphagocytic cells. A cis-element at −90 bp is required for derepressed transcription and serves as a binding site for multiple transcriptional activators. We now report that this element also serves as a binding site for CDP. The affinity of CDP for this element is relatively weak compared with upstream CDP-binding sites within the promoter, consistent with the promiscuous transcriptional activity exhibited by the −102 to +12 bp gp91phox promoter fragment. Further analysis of the proximal promoter reveals an additional weak-affinity CDP-binding site centered at approximately −20 bp. Overexpression of cloned CDP represses the −102 to +12 bp gp91phox promoter, indicating that these proximal CDP-binding sites are functionally significant. The constellation of transcriptional activators and a repressor that interacts with the −90 bp cis-element is identical to that observed for a promoter element at −220 bp, reflecting the highly modular organization of the gp91phoxpromoter. These studies illustrate the complex interplay between transcriptional activators and a repressor that contribute to the myeloid-restricted expression of the gp91phox gene.

scholarly journals Purification and identification of the STAT5 protease in myeloid cells

2007 ◽  
Vol 404 (1) ◽  
pp. 81-87 ◽  
Author(s):  
Björn Schuster ◽  
Lisa Hendry ◽  
Helen Byers ◽  
Steven F. Lynham ◽  
Malcolm A. Ward ◽  
...  
Keyword(s):  
Serine Protease ◽  
Active Form ◽  
Myeloid Cells ◽  
Specific Inhibitor ◽  
Myeloid Cell ◽  
Full Length ◽  
Cathepsin G ◽  
Transactivation Domain ◽  
Immature Myeloid Cell ◽  
Immature Myeloid Cells

STAT (signal transducer and activator of transcription) proteins are critical regulators of cytokine-induced cell proliferation, differentiation and survival. STAT functional activity can be variably regulated by post-translational modifications, including phosphorylation, acetylation, methylation and sumoylation. Additionally, limited proteolytic digestion of full-length STAT proteins (STATα) generates C-terminally truncated forms (STATγ) in different cell lineages, which have significantly reduced transcriptional activity due to the lack of the transactivation domain. Previously, it has been shown that STAT5γ, generated by an unidentified nuclear serine protease, plays an important role in myeloid cell differentiation and is aberrantly expressed in acute myeloid leukaemia. To better understand this regulatory mechanism for STAT5 function, we have purified the STAT5 protease from the immature myeloid cell line 32D and identified it by MS analysis as the granule-derived serine protease, CatG (cathepsin G). We show that purified CatG can specifically cleave full-length STAT5 to generate STAT5γ, and this activity can be inhibited by AEBSF [4-(2-aminoethyl)benzenesulfonyl fluoride] in an in vitro protease assay. Importantly, preparation of nuclear and cytoplasmic extracts from immature myeloid cell lines, 32D and FDC-P1, in the presence of a specific inhibitor for CatG results in the identification of STAT5α only. These studies indicate that nuclear STAT5γ does not naturally exist in immature myeloid cells and is artificially generated from STAT5α during the preparation of extracts due to the abundance of CatG in these cells. Therefore in contrast with earlier studies, our data suggest that STAT5α, rather than STAT5γ is the active form in immature myeloid cells.

scholarly journals Possible multiple binding sites for o-aminophenol on uridine diphosphate glucuronyltransferase

1977 ◽  
Vol 163 (1) ◽  
pp. 125-131 ◽  
Author(s):  
R D Howland ◽  
L D Bohm
Keyword(s):  
Binding Site ◽  
Binding Sites ◽  
Fold Increase ◽  
Prior Treatment ◽  
Uridine Diphosphate ◽  
Adult Rats ◽  
Ph Optimum ◽  
Acceptor Substrate ◽  
Multiple Binding Sites ◽  
Multiple Binding

1. Hepatic microsomal UDP-glucuronyltransferase (EC 2.4.1.17) derived from either weanling or adult rats exhibits three pH optima, at pH 5.4, 7.2 and 9.2, when o-aminophenol is the acceptor substrate, whereas p-nitrophenol is the acceptor substrate only on pH optimum is observed, at pH 5.4.2. Prior treatment of rats of either age with 3-methylcholanthrene results in a 2-3-fold increase in o-aminophenol conjugation at pH 5.4 and a 6-9-fold increase at pH 9.2. At pH 7.2, the induced enzyme is 2 to 3 times more active towards o-aminophenol than the control enzyme, but no pH optimum is demonstrable. 3. o-Aminophenol conjugation at pH 5.4 and 9.2 is inhibited competitively by both p-nitrophenol and p-nitrophenyl glucuronide, suggesting that the two phenolic aglycones share the same binding site. At pH 7.2, however, p-nitrophenyl glucuronide does not inhibit o-aminophenol conjugation, suggesting that the binding site at this pH is not shared by the two phenols. These data are consistent with the existence of more than one binding site for o-aminophenol on UDP-glucuronyltransferase.

scholarly journals PKCα and PKCδ Activation Regulates Transcriptional Activity and Degradation of Progesterone Receptor in Human Astrocytoma Cells

Endocrinology ◽  
2015 ◽  
Vol 156 (3) ◽  
pp. 1010-1022 ◽  
Author(s):  
Aliesha González-Arenas ◽  
Miguel Ángel Peña-Ortiz ◽  
Valeria Hansberg-Pastor ◽  
Brenda Marquina-Sánchez ◽  
Noemi Baranda-Ávila ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Transcriptional Activity ◽  
In Silico Analysis ◽  
Fold Increase ◽  
26S Proteasome ◽  
Astrocytoma Cells ◽  
Tetradecanoyl Phorbol Acetate ◽  
Human Astrocytoma ◽  
Subsequent Degradation ◽  
And Function

Abstract Progesterone regulates cancer cell proliferation and invasion through its receptors (PR-A and PR-B), whose phosphorylation modifies their transcriptional activity and induce their degradation. We identified by in silico analysis a putative residue (Ser400) in PR that might be phosphorylated by protein kinase C (PKC), a family of enzymes involved in the proliferation and infiltration of astrocytomas, the most frequent and aggressive brain tumors. A grade III human astrocytoma-derived cell line was used to study the role of PKC in PR phosphorylation, transcriptional activity, and degradation. Treatment with PKC activator [tetradecanoyl phorbol acetate (TPA)] increased PR phosphorylation in Ser400 after 5 minutes, which in turn induced PR transcriptional activity and its subsequent degradation by the 26S proteasome 3–5 hours after treatment. Silencing or inhibition of PKCα and PKCδ blocked PR phosphorylation and degradation induced by TPA. Both PR isoforms were associated with PKCα and reached the maximum association after 5 minutes of TPA addition. These data correlated with immunnofluorescence assays in which nuclear colocalization of PKCα with PR increased after TPA treatment. We observed a 2-fold increase in cell proliferation after PKC activation with TPA that was reduced with the PR antagonist, RU486. The PR S400A mutant revealed that this residue is essential for PKC-mediated PR phosphorylation and degradation. Our results show a key participation of PKCα and PKCδ in PR regulation and function.

scholarly journals Binding, sliding, and function of cohesin during transcriptional activation

2017 ◽  
Vol 114 (7) ◽  
pp. E1062-E1071 ◽  
Author(s):  
Melinda S. Borrie ◽  
John S. Campor ◽  
Hansa Joshi ◽  
Marc R. Gartenberg
Keyword(s):  
Transcriptional Activation ◽  
Binding Sites ◽  
Transcriptional Activity ◽  
Gene Activation ◽  
Gene Promoter ◽  
Chromatid Cohesion ◽  
M Phase ◽  
And Function ◽  
Transcriptional Induction ◽  
Dna Circularization

The ring-shaped cohesin complex orchestrates long-range DNA interactions to mediate sister chromatid cohesion and other aspects of chromosome structure and function. In the yeast Saccharomyces cerevisiae, the complex binds discrete sites along chromosomes, including positions within and around genes. Transcriptional activity redistributes the complex to the 3′ ends of convergently oriented gene pairs. Despite the wealth of information about where cohesin binds, little is known about cohesion at individual chromosomal binding sites and how transcription affects cohesion when cohesin complexes redistribute. In this study, we generated extrachromosomal DNA circles to study cohesion in response to transcriptional induction of a model gene, URA3. Functional cohesin complexes loaded onto the locus via a poly(dA:dT) tract in the gene promoter and mediated cohesion before induction. Upon transcription, the fate of these complexes depended on whether the DNA was circular or not. When gene activation occurred before DNA circularization, cohesion was lost. When activation occurred after DNA circularization, cohesion persisted. The presence of a convergently oriented gene also prevented transcription-driven loss of functional cohesin complexes, at least in M phase-arrested cells. The results are consistent with cohesin binding chromatin in a topological embrace and with transcription mobilizing functional complexes by sliding them along DNA.

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 Myb binding sites mediate negative regulation of c-myb expression in T- cell lines

Blood ◽  
1995 ◽  
Vol 86 (5) ◽  
pp. 1873-1880 ◽  
Author(s):  
J Guerra ◽  
DA Withers ◽  
LM Boxer
Keyword(s):  
T Cell ◽  
Binding Site ◽  
Cell Line ◽  
Cell Lines ◽  
Binding Sites ◽  
Myeloid Cell ◽  
Hematopoietic Cell ◽  
Wild Type ◽  
Myeloid Cell Line ◽  
T Cell Lines

In hematopoietic cell development, the c-myb transcription factor plays an important role. c-myb mRNA is expressed at high levels in immature proliferating cells and in leukemic cells. We have investigated the regulatory role of Myb protein binding to the human c-myb promoter. Three Myb binding sites have been described at approximately 600 bp upstream of the cap site. By transient transfection assays in hematopoietic cell lines, we found that deletion of the previously defined most 52 Myb binding site had no effect on activity, whereas deletion of the region containing the remaining two Myb binding sites resulted in an increase in activity in both a T-cell line and a myeloid cell line. To specifically test the importance of these two Myb binding sites, the activity of three-point mutation constructs was measured. Mutation of either Myb binding site resulted in an increase in activity compared with the wild-type promoter in T cells. Mutation of both sites produced even higher activity. Transfection of the Myb site mutants into the myeloid cell line resulted in no change in activity compared with the wild type construct. Results from gel shift analysis, UV cross- linking, and Western blots showed that both c-Myb and B-Myb bound to the Myb I and II sites. We conclude that the Myb family proteins negatively regulate c-myb expression in T-cell lines in contrast to the positive regulation via these sites, which has been shown in fibroblasts. In addition, in a myeloid cell line, the Myb binding sites are nonfunctional.

Liver-specific gene expression: A-activator-binding site, a promoter module present in vitellogenin and acute-phase genes

1991 ◽  
Vol 11 (1) ◽  
pp. 93-101
Author(s):  
M Kaling ◽  
W Kugler ◽  
K Ross ◽  
C Zoidl ◽  
G U Ryffel
Keyword(s):  
Transcription Factor ◽  
Binding Site ◽  
Acute Phase ◽  
Interleukin 6 ◽  
Binding Sites ◽  
Transcriptional Activity ◽  
In Vitro Transcription ◽  
Gel Retardation ◽  
Vitellogenin Gene

The A2 vitellogenin gene of Xenopus laevis, which is expressed liver specifically, contains an A-activator-binding site (AABS) that mediates high in vitro transcriptional activity in rat liver nuclear extracts. Footprint experiments with DNase I and gel retardation assays revealed the binding of several proteins to AABS. Using binding sites of known DNA-binding proteins as competitors in the gel retardation assay, we found that the transcription factor C/EBP and/or one of its "iso-binders" as well as LFB1/HNF1 bound AABS. These interactions were confirmed by in vitro transcription experiments using various oligonucleotides as competitors. However, saturating amounts of C/EBP- and LFB1/HNF1-binding sites as competitors only partially blocked AABS-mediated transcriptional activity. This finding implies that at least a third distinct transcription factor interacts with AABS. In vitro transcription experiments revealed that AABS was present not only in the closely related Xenopus A1 vitellogenin gene but also in acute-phase genes as a liver-specific regulatory element known to confer the interleukin-6 response. Both AABS and the interleukin-6 response element are promoter modules interacting with at least three distinct transcription factors, including C/EBP and LFB1/HNF1.

Sign in / Sign up

Export Citation Format

Share Document