FoxP3 maintains Treg unresponsiveness by selectively inhibiting the promoter DNA-binding activity of AP-1

Blood ◽  
2008 ◽  
Vol 111 (7) ◽  
pp. 3599-3606 ◽  
Author(s):  
Sang-Myeong Lee ◽  
Beixue Gao ◽  
Deyu Fang
Keyword(s):  
T Cells ◽  
Transcription Factors ◽  
Dna Binding ◽  
Transcriptional Activity ◽  
Molecular Mechanisms ◽  
Ectopic Expression ◽  
Binding Activity ◽  
Dna Binding Activity ◽  
N Terminus ◽  
Promoter Dna

Abstract Regulatory T cells (Tregs) have been shown to play a crucial role in maintaining self-tolerance and suppressing autoimmunity. The forkhead transcription factor, FoxP3, is a key molecule necessary and sufficient for Tregs development and function. However, the molecular mechanisms by which FoxP3 regulates the phenotypic (anergic) and the functional (suppressive) characteristics of Tregs are not well defined. Here we found that the promoter DNA-binding activity of AP-1 transcription factors is selectively inhibited in the naturally occurring CD4+ CD25+ Tregs from mice. The impaired AP-1 DNA binding is not the result of the decreased nuclear translocation of AP-1 family transcription factors, including c-Jun, JunB, and c-Fos. FoxP3 significantly suppresses both the transcriptional activity and promoter DNA-binding of AP-1 by interacting with c-Jun. The N-terminus of FoxP3, but not its C-terminus forkhead domain, specifically interacts with phosphorylated c-Jun and alters c-Jun subnuclear distribution. This N-terminus of FoxP3 with nuclear localization signals (FoxP3N/NLS) is able to suppress AP-1 transcriptional activity. Ectopic expression of FoxP3N/NLS sufficiently induces the unresponsiveness of mouse primary CD4+ CD25− T cells, whereas the full-length FoxP3 is required for the suppressive functions of Tregs. These findings uncover one of the mechanisms underlying how FoxP3 maintains the unresponsiveness of Tregs.

Single-Stranded DNA-Binding Proteins (SSBPs) Regulate the Abundance of the LIM-Homeodomain Protein LHX2 and Augment Its Transcriptional Activity.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1239-1239
Author(s):  
Ying Cai ◽  
Zhixiong Xu ◽  
Lalitha Nagarajan ◽  
Stephen J. Brandt
Keyword(s):  
Transcription Factors ◽  
Dna Binding ◽  
Protein Turnover ◽  
Transcriptional Activity ◽  
Binding Activity ◽  
Protein Abundance ◽  
Single Stranded Dna ◽  
Dna Binding Activity ◽  
E Box ◽  
Lim Homeodomain

Abstract A small family of proteins with putative single-stranded DNA-binding activity has been shown to augment the biological actions of LIM-homeodomain (LIM-HD) transcription factors through the mediation of the LIM domain-binding protein LDB1. We recently established that two of these SSBPs, Ssbp2 and Ssbp3, were components of an E-box-GATA DNA-binding complex in murine erythroid progenitors containing transcription factors Tal1, E2A, and Gata-1 and LIM-only protein Lmo2 and showed that Ssbp2 stimulated E box-GATA DNA-binding activity by inhibiting Ldb1 ubiquitination and Ldb1 and Lmo2 turnover (Genes & Dev.21:942–955, 2007). Since LIM-HD proteins are substrates of different E3 ubiquitin ligases than LIM-only proteins and have the additional property of binding DNA, we sought to determine the effect of SSBPs on LIM-HD expression and function. Using the prototype LIM-HD protein Lhx2 and one of its best-characterized target genes, Cga, for analysis, we found that an Ssbp3-, Ldb1-, and Lhx2-containing complex associated with an Lhx2 binding element in the Cga promoter in vitro and in mouse pituitary cells (alphaT3-1 cell line) in vivo. We then showed that enforced expression of Ssbp2 and Ssbp3 in alphaT3-1 cells increased Lhx2 and Ldb1 protein abundance, Lhx2 DNA-binding activity, and Cga expression and augmented Lhx2 transcriptional activity in an Ldb1-dependent fashion. While Lhx2-Ldb1-Ssbp3 DNA-binding activity increased in Ssbp3- relative to vector-transfected cells, the affinity of this complex for DNA was unaltered. Similar to the effect of Ssbp2 on Lmo2 in murine erythroleukemia (MEL) cells, overexpressed Ssbp3 reduced Lhx2 protein turnover in cycloheximide-treated alphaT3-1 cells without affecting Lhx2 RNA levels. In contrast, knockdown of endogenous Ssbp3, but not Ssbp2 which is expressed at much lower levels in these cells, reduced Lhx2 and Ldb1 abundance, Lhx2 DNA-binding activity, Lhx2, Ldb1, and Ssbp3 loading onto the Cga promoter, Cga promoter activity, and endogenous Cga gene expression. Significantly, neither overexpression nor knockdown of Ssbp2 in MEL cells, which express both the LIM-only protein Lmo2 and LIM-HD protein Lhx2, affected Lhx2 protein abundance, and Lhx2 DNA-binding activity was undetectable in nuclear extracts from these cells despite the presence of immunoreactive Lhx2. These studies indicate that SSBP augmentation of LIM-HD function results from Ldb1-mediated inhibition of LIM-HD protein turnover and increased assembly of a LIM-HD/LDB1/SSBP DNA-binding complex. The much greater affinity for LDB1 of LIM-only compared to LIM-HD proteins is likely a major determinant of the SSBP effect on LIM-HD protein abundance. Finally, these findings are consistent with cell type-specific contributions of different SSBPs, even for similar LDB1-dependent actions.

scholarly journals Chasing the structural diversity of the transcription regulator Mycobacterium tuberculosis HigA2

IUCrJ ◽  
2021 ◽  
Vol 8 (5) ◽  
Author(s):  
William Richardson ◽  
Gyun Won Kang ◽  
Hee Joong Lee ◽  
Kang Mu Kwon ◽  
Saron Kim ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Transcription Factors ◽  
Dna Binding ◽  
Dna Sequences ◽  
Molecular Mechanisms ◽  
Structural Diversity ◽  
Structural Rearrangement ◽  
Binding Activity ◽  
Dna Binding Activity ◽  
Α Helix

Transcription factors are the primary regulators of gene expression and recognize specific DNA sequences under diverse physiological conditions. Although they are vital for many important cellular processes, it remains unclear when and how transcription factors and DNA interact. The antitoxin from a toxin–antitoxin system is an example of negative transcriptional autoregulation: during expression of the cognate toxin it is suppressed through binding to a specific DNA sequence. In the present study, the antitoxin HigA2 from Mycobacterium tuberculosis M37Rv was structurally examined. The crystal structure of M. tuberculosis HigA2 comprises three sections: an N-terminal autocleavage region, an α-helix bundle which contains an HTH motif, and a C-terminal β-lid. The N-terminal region is responsible for toxin binding, but was shown to cleave spontaneously in its absence. The HTH motif performs a key role in DNA binding, with the C-terminal β-lid influencing the interaction by mediating the distance between the motifs. However, M. tuberculosis HigA2 exhibits a unique coordination of the HTH motif and no DNA-binding activity is detected. Three crystal structures of M. tuberculosis HigA2 show a flexible alignment of the HTH motif, which implies that the motif undergoes structural rearrangement to interact with DNA. This study reveals the molecular mechanisms of how transcription factors interact with partner proteins or DNA.

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.

Differential use of SCL/TAL-1 DNA-binding domain in developmental hematopoiesis

Blood ◽  
2008 ◽  
Vol 112 (4) ◽  
pp. 1056-1067 ◽  
Author(s):  
Mira T. Kassouf ◽  
Hedia Chagraoui ◽  
Paresh Vyas ◽  
Catherine Porcher
Keyword(s):  
Dna Binding ◽  
Molecular Mechanisms ◽  
Binding Activity ◽  
Hematopoietic Stem ◽  
Helix Loop Helix ◽  
Experimental Systems ◽  
Dna Binding Activity ◽  
Independent Functions

Abstract Dissecting the molecular mechanisms used by developmental regulators is essential to understand tissue specification/differentiation. SCL/TAL-1 is a basic helix-loop-helix transcription factor absolutely critical for hematopoietic stem/progenitor cell specification and lineage maturation. Using in vitro and forced expression experimental systems, we previously suggested that SCL might have DNA-binding–independent functions. Here, to assess the requirements for SCL DNA-binding activity in vivo, we examined hematopoietic development in mice carrying a germline DNA-binding mutation. Remarkably, in contrast to complete absence of hematopoiesis and early lethality in scl-null embryos, specification of hematopoietic cells occurred in homozygous mutant embryos, indicating that direct DNA binding is dispensable for this process. Lethality was forestalled to later in development, although some mice survived to adulthood. Anemia was documented throughout development and in adulthood. Cellular and molecular studies showed requirements for SCL direct DNA binding in red cell maturation and indicated that scl expression is positively autoregulated in terminally differentiating erythroid cells. Thus, different mechanisms of SCL's action predominate depending on the developmental/cellular context: indirect DNA binding activities and/or sequestration of other nuclear regulators are sufficient in specification processes, whereas direct DNA binding functions with transcriptional autoregulation are critically required in terminal maturation processes.

Human T-cell lymphotropic virus oncoprotein Tax represses TGF-β1 signaling in human T cells via c-Jun activation: a potential mechanism of HTLV-I leukemogenesis

Blood ◽  
2002 ◽  
Vol 100 (12) ◽  
pp. 4129-4138 ◽  
Author(s):  
Bertrand Arnulf ◽  
Aude Villemain ◽  
Christophe Nicot ◽  
Elodie Mordelet ◽  
Pierre Charneau ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Dna Binding ◽  
Binding Activity ◽  
T Cell Leukemia ◽  
Cell Leukemia ◽  
Dna Binding Activity ◽  
Human T Cell ◽  
Peripheral T Cells ◽  
Tgf Β1

Human T-cell leukemia virus I is the etiologic agent of adult T-cell leukemia (ATL), an aggressive T-cell malignancy. The viral oncoprotein Tax, through the activation of nuclear factorκB (NF-κB), CCAAT-enhancer binding protein (CREB), and activated protein-1 (AP-1) pathways, is a transcriptional regulator of critical genes for T-cell homeostasis. In ATL cells, activated AP-1 complexes induce the production of transforming growth factor β1 (TGF-β1). TGF-β1 is an inhibitor of T-cell proliferation and cytotoxicity. Here we show that, in contrast to normal peripheral T cells, ATL cells are resistant to TGF-β1–induced growth inhibition. The retroviral transduction of the Tax protein in peripheral T cells resulted in the loss of TGF-β1 sensitivity. Transient transfection of Tax in HepG2 cells specifically inhibited Smad/TGF-β1 signaling in a dose-dependent manner. In the presence of Tax transfection, increasing amounts of Smad3 restored TGF-β1 signaling. Tax mutants unable to activate NF-κB or CREB pathways were also able to repress Smad3 transcriptional activity. Next we have demonstrated that Tax inhibits TGF-β1 signaling by reducing the Smad3 DNA binding activity. However, Tax did not decrease the expression and the nuclear translocation of Smad3 nor did it interact physically with Smad3. Rather, Tax induced c-Jun N-terminal kinase (JNK) activity and c-Jun phosphorylation, leading to the formation of Smad3/c-Jun complexes. Whereas c-Jun alone abrogates Smad3 DNA binding, cotransfection of Tax and of a dominant-negative form of JNK or a c-Jun antisense-restored Smad3 DNA binding activity and TGF-β1 responsiveness. In ATL and in normal T cells transduced by Tax, c-Jun was constitutively phosphorylated. Thus, we describe a new function of Tax, as a repressor of TGF-β1 signaling through JNK/c-Jun constitutive activation, which may play a critical role in ATL leukemogenesis.

scholarly journals Drosophila Mi-2 Negatively Regulates dDREF by Inhibiting Its DNA-Binding Activity

2002 ◽  
Vol 22 (14) ◽  
pp. 5182-5193 ◽  
Author(s):  
Fumiko Hirose ◽  
Nobuko Ohshima ◽  
Eun-Jeong Kwon ◽  
Hideki Yoshida ◽  
Masamitsu Yamaguchi
Keyword(s):  
Dna Binding ◽  
Polytene Chromosomes ◽  
Ectopic Expression ◽  
Binding Activity ◽  
Mobility Shift ◽  
Interacting Protein ◽  
Reciprocal Regulation ◽  
Dna Binding Activity ◽  
Expression Of Genes ◽  
Biochemical Analyses

ABSTRACT Drosophila melanogaster DNA replication-related element (DRE) factor (dDREF) is a transcriptional regulatory factor required for the expression of genes carrying the 5′-TATCGATA DRE. dDREF has been reported to bind to a sequence in the chromatin boundary element, and thus, dDREF may play a part in regulating insulator activity. To generate further insights into dDREF function, we carried out a Saccharomyces cerevisiae two-hybrid screening with DREF polypeptide as bait and identified Mi-2 as a DREF-interacting protein. Biochemical analyses revealed that the C-terminal region of Drosophila Mi-2 (dMi-2) specifically binds to the DNA-binding domain of dDREF. Electrophoretic mobility shift assays showed that dMi-2 thereby inhibits the DNA-binding activity of dDREF. Ectopic expression of dDREF and dMi-2 in eye imaginal discs resulted in severe and mild rough-eye phenotypes, respectively, whereas flies simultaneously expressing both proteins exhibited almost-normal eye phenotypes. Half-dose reduction of the dMi-2 gene enhanced the DREF-induced rough-eye phenotype. Immunostaining of polytene chromosomes of salivary glands showed that dDREF and dMi-2 bind in mutually exclusive ways. These lines of evidence define a novel function of dMi-2 in the negative regulation of dDREF by its DNA-binding activity. Finally, we postulated that dDREF and dMi-2 may demonstrate reciprocal regulation of their functions.

scholarly journals Direct inhibition of the DNA-binding activity of POU transcription factors Pit-1 and Brn-3 by selective binding of a phenyl-furan-benzimidazole dication

2008 ◽  
Vol 36 (10) ◽  
pp. 3341-3353 ◽  
Author(s):  
Paul Peixoto ◽  
Yang Liu ◽  
Sabine Depauw ◽  
Marie-Paule Hildebrand ◽  
David W. Boykin ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Dna Binding ◽  
Binding Activity ◽  
Direct Inhibition ◽  
Selective Binding ◽  
Dna Binding Activity

scholarly journals DNA Binding Activity of the Herpes Simplex Virus Type 1 Origin Binding Protein, UL9, Can Be Modulated by Sequences in the N Terminus: Correlation between Transdominance and DNA Binding

2006 ◽  
Vol 80 (9) ◽  
pp. 4491-4500 ◽  
Author(s):  
Soma Chattopadhyay ◽  
Sandra K. Weller
Keyword(s):  
Herpes Simplex ◽  
Dna Binding ◽  
Binding Activity ◽  
Plaque Formation ◽  
Dna Binding Domain ◽  
Dna Binding Activity ◽  
Origin Binding Protein ◽  
N Terminus ◽  
Simplex Virus

ABSTRACT UL9, the origin binding protein of herpes simplex virus type 1, is a member of the SF2 family of helicases. Cotransfection of cells with infectious viral DNA and plasmids expressing either full-length UL9 or the C-terminal DNA binding domain alone results in the drastic inhibition of plaque formation which can be partially relieved by an insertion mutant lacking DNA binding activity. In this work, C-terminally truncated mutants which terminate at or near residue 359 were shown to potentiate plaque formation, while other C-terminal truncations were inhibitory. Thus, residues in the N-terminal region appear to regulate the inhibitory properties of UL9. To identify which residues were involved in this regulation, a series of N-terminally truncated mutants were constructed which contain the DNA binding domain and various N-terminal extensions. Mutants whose N terminus is either at residue 494 or 535 were able to bind the origin efficiently and were inhibitory to plaque formation, whereas constructs whose N terminus is at residue 304 or 394 were defective in origin binding activity and were able to relieve inhibition. Since UL9 is required for viral infection at early but not late times and is inhibitory to infection when overexpressed, we propose that the DNA binding activities of UL9 are regulated during infection. For infection to proceed, UL9 may need to switch from a DNA binding to a non-DNA binding mode, and we suggest that sequences residing in the N terminus play a role in this switch.

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