scholarly journals Transcription Factor SCL Is Required for c-kit Expression and c-Kit Function in Hemopoietic Cells

1998 ◽  
Vol 188 (3) ◽  
pp. 439-450 ◽  
Author(s):  
Gorazd Krosl ◽  
Gang He ◽  
Martin Lefrancois ◽  
Frédéric Charron ◽  
Paul-Henri Roméo ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Cell Survival ◽  
Cell Line ◽  
Ectopic Expression ◽  
Binding Activity ◽  
Dominant Negative ◽  
Hemopoietic Cells ◽  
Downstream Target ◽  
Helix Loop Helix ◽  
Dna Binding Activity

In normal hemopoietic cells that are dependent on specific growth factors for cell survival, the expression of the basic helix-loop-helix transcription factor SCL/Tal1 correlates with that of c-Kit, the receptor for Steel factor (SF) or stem cell factor. To address the possibility that SCL may function upstream of c-kit, we sought to modulate endogenous SCL function in the CD34+ hemopoietic cell line TF-1, which requires SF, granulocyte/macrophage colony–stimulating factor, or interleukin 3 for survival. Ectopic expression of an antisense SCL cDNA (as-SCL) or a dominant negative SCL (dn-SCL) in these cells impaired SCL DNA binding activity, and prevented the suppression of apoptosis by SF only, indicating that SCL is required for c-Kit–dependent cell survival. Consistent with the lack of response to SF, the level of c-kit mRNA and c-Kit protein was significantly and specifically reduced in as-SCL– or dn-SCL– expressing cells. c-kit mRNA, c-kit promoter activity, and the response to SF were rescued by SCL overexpression in the antisense or dn-SCL transfectants. Furthermore, ectopic c-kit expression in as-SCL transfectants is sufficient to restore cell survival in response to SF. Finally, enforced SCL in the pro–B cell line Ba/F3, which is both SCL and c-kit negative is sufficient to induce c-Kit and SF responsiveness. Together, these results indicate that c-kit, a gene that is essential for the survival of primitive hemopoietic cells, is a downstream target of the transcription factor SCL.

Treatment of cultured myotubes with the proteasome inhibitor β-lactone increases the expression of the transcription factor C/EBPβ

2007 ◽  
Vol 292 (1) ◽  
pp. C216-C226 ◽  
Author(s):  
Wei Wei ◽  
Hongmei Yang ◽  
Michael Menconi ◽  
Peirang Cao ◽  
Chester E. Chamberlain ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Transcription Factor ◽  
Dna Binding ◽  
Proteasome Inhibitor ◽  
Binding Activity ◽  
Dominant Negative ◽  
Dna Binding Activity ◽  
Cultured Myotubes ◽  
Nuclear Levels ◽  
Factor C

The role of the proteasome in the regulation of cellular levels of the transcription factor CCAAT/enhancer-binding protein β (C/EBPβ) is poorly understood. We tested the hypothesis that C/EBPβ levels in cultured myotubes are regulated, at least in part, by proteasome activity. Treatment of cultured L6 myotubes, a rat skeletal muscle cell line, with the specific proteasome inhibitor β-lactone resulted in increased nuclear levels of C/EBPβ as determined by Western blotting and immunofluorescent detection. This effect of β-lactone reflected inhibited degradation of C/EBPβ. Surprisingly, the increased C/EBPβ levels in β-lactone-treated myotubes did not result in increased DNA-binding activity. In additional experiments, treatment of the myotubes with β-lactone resulted in increased nuclear levels of growth arrest DNA damage/C/EBP homologous protein (Gadd153/CHOP), a dominant-negative member of the C/EBP family that can form heterodimers with other members of the C/EBP family and block DNA binding. Coimmunoprecipitation and immunofluorescent detection provided evidence that C/EBPβ and Gadd153/CHOP interacted and colocalized in the nuclei of the β-lactone-treated myotubes. When Gadd153/CHOP expression was downregulated by transfection of myotubes with siRNA targeting Gadd153/CHOP, C/EBPβ DNA-binding activity was restored in β-lactone-treated myotubes. The results suggest that C/EBPβ is degraded by a proteasome-dependent mechanism in skeletal muscle cells and that Gadd153/CHOP can interact with C/EBPβ and block its DNA-binding activity. The observations are important because they increase the understanding of the complex regulation of the expression and activity of C/EBPβ in skeletal muscle.

Over-Expression of the P65 Subunit of NF-kB and Increased DNA Binding Activity of NF-kB Is a Common Event Both in Philadelphia Positive and Negative Chronic Myeloproliferative Disorders.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4759-4759
Author(s):  
Alessandro Morotti ◽  
Veronica Ullmannova ◽  
Daniela Cilloni ◽  
Francesca Messa ◽  
Manuela Messa ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Transcription Factor ◽  
Cell Growth ◽  
Dna Binding ◽  
Western Blot ◽  
Cell Line ◽  
Myeloproliferative Disorders ◽  
Binding Activity ◽  
K562 Cell Line ◽  
Dna Binding Activity

Abstract The chronic myeloproliferative diseases (CMPD) are clonal disorders characterized by increased proliferation of cells from one or more myeloid lineages. The most common CMPD is Chronic Myeloid Leukemia which is characterized by the Philadelphia t(9;22) chromosomal translocation. The pathogenesis of Philadelphia negative CMPD is poorly understood, although the activation of tyrosine kinases appears to be an essential feature. For example, a constitutively activated PDGF receptor tyrosine kinase (FIP1L1-PDGFRA) is involved in some cases of the hypereosinophilic syndrome (HES), which is a form of CMPD characterized by increased proliferation of eosinophils. Different reports have demonstrated that the transcription factor NF-kB is essential for Bcr-Abl mediated transformation. NF-kB is a transcription factor which is composed of two subunits (generally p65 and p50). NF-kB dimers are retained into the cytoplasm by the inhibitory protein IkB. Different stimuli trigger the Serine phosphorylation of IkB and its proteolitc degradation. Free NF-kB translocates into the nucleus where it mediates the transcription of different genes involved in cellular proliferation, transformation and in apoptosis resistance. The aim of this work is to evaluate whether NF-kB is active both in Ph positive and in Ph negative CMPD. Bone marrow samples of 8 myeloproliferative disorders (3 Philadelphia positive CML, 3 Ph negative CML-like, 1 HES, 1 Idiopathic Myelofibrosis) have been collected at the diagnosis. The t(9;22) positive K562 cell line, derived form a CML blast crysis, has been used as a positive cellular control of the following experiments. Each samples have been lysed to obtain cytosolic and nuclear extracts. Western blot have been performed to evaluate the expression of the p65 subunit of NF-kB, the regulatory protein IkB and the antiapoptotic protein Bcl-2, whose expression may be regulated by NF-kB. Subsequently the DNA binding activity of NF-kB have been measured with an ELISA method. Our data shows that in all samples p65 is over-expressed both in the cytosol and in the nucleus respect to normal peripheral blood and normal bone marrow samples. The antiapoptotic Bcl-2 is also detectable by western blot in all pathological samples. In normal samples IkB is detected only in the cytosol and not in the nucleus while in CMPD samples it is expressed predominately in the nucleus. Basal DNA binding activity of NF-kB is increased in all the nuclear samples but not in normal samples. To assess whether NF-kB is directly involved in the control of cell growth and apoptosis, we have developed a stable K562 cell line expressing the super repressor IkB. The SR-IkB is a mutated for of IkB which can not be degraded causing an cytosolic sequestration of NF-kB. This stable cell line has a marked reduction of cell growth and is more sensible to the apoptotic stimuli. The data described above suggest that NF-kB activation may be a common mechanism of transformation in both Ph positive and negative CMPD and that its inhibition may be a powerful targeted molecular therapy.

scholarly journals Silencing of the inhibitor of DNA binding protein 4 (ID4) contributes to the pathogenesis of mouse and human CLL

Blood ◽  
2011 ◽  
Vol 117 (3) ◽  
pp. 862-871 ◽  
Author(s):  
Shih-Shih Chen ◽  
Rainer Claus ◽  
David M. Lucas ◽  
Lianbo Yu ◽  
Jiang Qian ◽  
...  
Keyword(s):  
Dna Binding ◽  
Promoter Methylation ◽  
Binding Protein ◽  
Dna Binding Protein ◽  
Binding Activity ◽  
Lymphocytic Leukemia ◽  
Dominant Negative ◽  
Helix Loop Helix ◽  
Dna Binding Activity ◽  
Inhibitor Of Dna Binding

Abstract Inhibitor of DNA binding protein 4 (ID4) is a member of the dominant-negative basic helix-loop-helix transcription factor family that lacks DNA binding activity and has tumor suppressor function. ID4 promoter methylation has been reported in acute myeloid leukemia and chronic lymphocytic leukemia (CLL), although the expression, function, and clinical relevance of this gene have not been characterized in either disease. We demonstrate that the promoter of ID4 is consistently methylated to various degrees in CLL cells, and increased promoter methylation in a univariable analysis correlates with shortened patient survival. However, ID4 mRNA and protein expression is uniformly silenced in CLL cells irrespective of the degree of promoter methylation. The crossing of ID4+/− mice with Eμ-TCL1 mice triggers a more aggressive murine CLL as measured by lymphocyte count and inferior survival. Hemizygous loss of ID4 in nontransformed TCL1-positive B cells enhances cell proliferation triggered by CpG oligonucleotides and decreases sensitivity to dexamethasone-mediated apoptosis. Collectively, this study confirms the importance of the silencing of ID4 in murine and human CLL pathogenesis.

The CIB1 Transcription Factor Regulates Light- and Heat-inducible Cell Elongation via a Two-step HLH/bHLH System

2020 ◽  
Author(s):  
Miho Ikeda ◽  
Nobutaka Mitsuda ◽  
Toru Ishizuka ◽  
Mai Satoh ◽  
Masaru Ohme-Takagi
Keyword(s):  
Transcription Factor ◽  
High Temperature ◽  
Dna Binding ◽  
Cell Elongation ◽  
Binding Activity ◽  
Bhlh Transcription Factor ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
Dna Binding Activity ◽  
Underlying Mechanisms

Abstract Light and high temperature promote plant cell elongation. PHYTOCHROME INTERACTING FACTOR4 (PIF4, a typical basic helix-loop-helix [bHLH] transcriptional activator) and the non-DNA-binding atypical HLH inhibitors PHYTOCHROME RAPIDLY REGULATED1 (PAR1) and LONG HYPOCOTYL IN FAR-RED 1 (HFR1) competitively regulate cell elongation in response to light conditions and high temperature. However, the underlying mechanisms have not been fully clarified. Here, we show that in Arabidopsis thaliana, the bHLH transcription factor CRYPTOCHROME-INTERACTING BASIC HELIX-LOOP-HELIX 1 (CIB1) positively regulates cell elongation under the control of PIF4, PAR1, and HFR1. Furthermore, PIF4 directly regulates CIB1 expression by interacting with its promoter, and PAR1 and HFR1 interfere with PIF4 binding to the CIB1 promoter. CIB1 activates genes that function in cell elongation, and PAR1 interferes with the DNA binding activity of CIB1, thus suppressing cell elongation. Hence, two antagonistic HLH/bHLH systems, the PIF4–PAR1/HFR1 and CIB1–PAR1 systems, regulate cell elongation in response to light and high temperature. We thus demonstrate the important role of non-DNA-binding small HLH proteins in the transcriptional regulation of cell elongation in plants.

Mechanistic role of heat shock protein 70 in Bcr-Abl–mediated resistance to apoptosis in human acute leukemia cells

Blood ◽  
2005 ◽  
Vol 105 (3) ◽  
pp. 1246-1255 ◽  
Author(s):  
Fei Guo ◽  
Celia Sigua ◽  
Purva Bali ◽  
Prince George ◽  
Warren Fiskus ◽  
...  
Keyword(s):  
Heat Shock ◽  
Heat Shock Protein ◽  
Myeloid Leukemia ◽  
Blast Crisis ◽  
Ectopic Expression ◽  
Binding Activity ◽  
Dominant Negative ◽  
Leukemia Cells ◽  
Dna Binding Activity ◽  
Induced Apoptosis

AbstractBcr-Abl–expressing primary or cultured leukemia cells display high levels of the antiapoptotic heat shock protein (hsp) 70 and are resistant to cytarabine (Ara-C), etoposide, or Apo-2L/TRAIL (TNF-related apoptosis-inducing ligand)–induced apoptosis. Conversely, a stable expression of the cDNA of hsp70 in the reverse orientation attenuated not only hsp70 but also signal transducers and activators of transcription 5 (STAT5) and Bcl-xL levels. This increased apoptosis induced by cytarabine, etoposide, or Apo-2L/TRAIL. Ectopic expression of hsp70 in HL-60 cells (HL-60/hsp70) inhibited Ara-C and etoposide-induced Bax conformation change and translocation to the mitochondria; attenuated the accumulation of cytochrome c, Smac, and Omi/HtrA2 in the cytosol; and inhibited the processing and activity of caspase-9 and caspase-3. Hsp70 was bound to death receptors 4 and 5 (DR4 and DR5) and inhibited Apo-2L/TRAIL-induced assembly and activity of the death-inducing signaling complex (DISC). HL-60/hsp70 cells exhibited increased levels and DNA binding activity of STAT5, which was associated with high levels of Pim-2 and Bcl-xL and resistance to apoptosis. Expression of the dominant negative (DN) STAT5 resensitized HL-60/hsp70 cells to cytarabine, etoposide, and Apo-2L/TRAIL–induced apoptosis. Collectively, these findings suggest that hsp70 inhibits apoptosis upstream and downstream of the mitochondria and is a promising therapeutic target for reversing drug-resistance in chronic myeloid leukemia-blast crisis and acute myeloid leukemia cells. (Blood. 2005;105:1246-1255)

A transcriptional repressive role for epithelial-specific ETS factor ELF3 on oestrogen receptor alpha in breast cancer cells

2016 ◽  
Vol 473 (8) ◽  
pp. 1047-1061 ◽  
Author(s):  
Vijaya Narasihma Reddy Gajulapalli ◽  
Venkata Subramanyam Kumar Samanthapudi ◽  
Madhusudana Pulaganti ◽  
Saratchandra Singh Khumukcham ◽  
Vijaya Lakhsmi Malisetty ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Transcription Factor ◽  
Dna Binding ◽  
Cancer Cells ◽  
Oestrogen Receptor ◽  
Breast Cancer Cells ◽  
Transcriptional Activity ◽  
Ectopic Expression ◽  
Binding Activity ◽  
Dna Binding Activity

Oestrogen receptor-α (ERα) is a ligand-dependent transcription factor that primarily mediates oestrogen (E2)-dependent gene transcription required for mammary gland development. Coregulators critically regulate ERα transcription functions by directly interacting with it. In the present study, we report that ELF3, an epithelial-specific ETS transcription factor, acts as a transcriptional repressor of ERα. Co-immunoprecipitation (Co-IP) analysis demonstrated that ELF3 strongly binds to ERα in the absence of E2, but ELF3 dissociation occurs upon E2 treatment in a dose- and time-dependent manner suggesting that E2 negatively influences such interaction. Domain mapping studies further revealed that the ETS (E-twenty six) domain of ELF3 interacts with the DNA binding domain of ERα. Accordingly, ELF3 inhibited ERα’s DNA binding activity by preventing receptor dimerization, partly explaining the mechanism by which ELF3 represses ERα transcriptional activity. Ectopic expression of ELF3 decreases ERα transcriptional activity as demonstrated by oestrogen response elements (ERE)-luciferase reporter assay or by endogenous ERα target genes. Conversely ELF3 knockdown increases ERα transcriptional activity. Consistent with these results, ELF3 ectopic expression decreases E2-dependent MCF7 cell proliferation whereas ELF3 knockdown increases it. We also found that E2 induces ELF3 expression in MCF7 cells suggesting a negative feedback regulation of ERα signalling in breast cancer cells. A small peptide sequence of ELF3 derived through functional interaction between ERα and ELF3 could inhibit DNA binding activity of ERα and breast cancer cell growth. These findings demonstrate that ELF3 is a novel transcriptional repressor of ERα in breast cancer cells. Peptide interaction studies further represent a novel therapeutic option in breast cancer therapy.

scholarly journals Mutations in the DNA-binding Domain of the Transcription Factor Bright Act as Dominant Negative Proteins and Interfere with Immunoglobulin Transactivation

2004 ◽  
Vol 279 (50) ◽  
pp. 52465-52472 ◽  
Author(s):  
Jamee C. Nixon ◽  
Jaya Rajaiya ◽  
Carol F. Webb
Keyword(s):  
Transcription Factor ◽  
Dna Binding ◽  
B Cell ◽  
Heavy Chain ◽  
Cell Development ◽  
Binding Activity ◽  
B Cell Development ◽  
Dominant Negative ◽  
Wild Type ◽  
Dna Binding Activity

Bright, for B cell regulator of immunoglobulin heavy chain transcription, binds A+T-rich sequences in the intronic enhancer regions of the murine heavy chain locus and 5′-flanking sequences of some variable heavy chain promoters. Most resting B cells do not express Bright; however, it is induced after stimulation with antigen or polyclonal mitogens. Bright activation results in up-regulation of μ transcription; however, it is not clear whether Bright function is critical for normal B cell development. To begin to address Bright function during B cell development, seven mutated forms of Bright were produced. Five of the seven mutants revealed little or no DNA binding activity. Furthermore, because Bright binds DNA as a dimer, two of the mutants formed complexes with wild type Bright and acted in a dominant negative fashion. Dominant negative Bright prevented the up-regulation of μ transcription in transfected Chinese hamster ovary cells transfected with wild type Bright. These data identify regions within Bright that are required for the DNA binding activity of Bright and for its function as a transcription factor.

Sign in / Sign up

Export Citation Format

Share Document