scholarly journals Virus-Dependent Phosphorylation of the IRF-3 Transcription Factor Regulates Nuclear Translocation, Transactivation Potential, and Proteasome-Mediated Degradation

1998 ◽  
Vol 18 (5) ◽  
pp. 2986-2996 ◽  
Author(s):  
Rongtuan Lin ◽  
Christophe Heylbroeck ◽  
Paula M. Pitha ◽  
John Hiscott
Keyword(s):  
Virus Infection ◽  
Transcriptional Activation ◽  
Nuclear Translocation ◽  
Sendai Virus ◽  
Active Form ◽  
Proteasome Inhibitors ◽  
Point Mutations ◽  
Regulatory Elements ◽  
Gene Promoters ◽  
Creb Binding Protein

ABSTRACT The interferon regulatory factors (IRF) consist of a growing family of related transcription proteins first identified as regulators of the alpha beta interferon (IFN-α/β) gene promoters, as well as the interferon-stimulated response element (ISRE) of some IFN-stimulated genes. IRF-3 was originally identified as a member of the IRF family based on homology with other IRF family members and on binding to the ISRE of the ISG15 promoter. IRF-3 is expressed constitutively in a variety of tissues, and the relative levels of IRF-3 mRNA do not change in virus-infected or IFN-treated cells. In the present study, we demonstrate that following Sendai virus infection, IRF-3 is posttranslationally modified by protein phosphorylation at multiple serine and threonine residues, which are located in the carboxy terminus of IRF-3. A combination of IRF-3 deletion and point mutations localized the inducible phosphorylation sites to the region -ISNSHPLSLTSDQ- between amino acids 395 and 407; point mutation of residues Ser-396 and Ser-398 eliminated virus-induced phosphorylation of IRF-3 protein, although residues Ser-402, Thr-404, and Ser-405 were also targets. Phosphorylation results in the cytoplasm-to-nucleus translocation of IRF-3, DNA binding, and increased transcriptional activation. Substitution of the Ser-Thr sites with the phosphomimetic Asp generated a constitutively active form of IRF-3 that functioned as a very strong activator of promoters containing PRDI-PRDIII or ISRE regulatory elements. Phosphorylation also appears to represent a signal for virus-mediated degradation, since the virus-induced turnover of IRF-3 was prevented by mutation of the IRF-3 Ser-Thr cluster or by proteasome inhibitors. Interestingly, virus infection resulted in the association of IRF-3 with the CREB binding protein (CBP) coactivator, as detected by coimmunoprecipitation with anti-CBP antibody, an interaction mediated by the C-terminal domains of both proteins. Mutation of residues Ser-396 and Ser-398 in IRF-3 abrogated its binding to CBP. These results are discussed in terms of a model in which virus-inducible, C-terminal phosphorylation of IRF-3 alters protein conformation to permit nuclear translocation, association with transcriptional partners, and primary activation of IFN- and IFN-responsive genes.

scholarly journals The Ebola Virus VP35 Protein Inhibits Activation of Interferon Regulatory Factor 3

2003 ◽  
Vol 77 (14) ◽  
pp. 7945-7956 ◽  
Author(s):  
Christopher F. Basler ◽  
Andrea Mikulasova ◽  
Luis Martinez-Sobrido ◽  
Jason Paragas ◽  
Elke Mühlberger ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Virus Infection ◽  
Transcriptional Activation ◽  
Ebola Virus ◽  
Nuclear Translocation ◽  
Sendai Virus ◽  
Vero Cells ◽  
Nuclear Accumulation ◽  
Regulatory Factor ◽  
Cellular Transcription Factor

ABSTRACT The Ebola virus VP35 protein was previously found to act as an interferon (IFN) antagonist which could complement growth of influenza delNS1 virus, a mutant influenza virus lacking the influenza virus IFN antagonist protein, NS1. The Ebola virus VP35 could also prevent the virus- or double-stranded RNA-mediated transcriptional activation of both the beta IFN (IFN-β) promoter and the IFN-stimulated ISG54 promoter (C. Basler et al., Proc. Natl. Acad. Sci. USA 97:12289-12294, 2000). We now show that VP35 inhibits virus infection-induced transcriptional activation of IFN regulatory factor 3 (IRF-3)-responsive mammalian promoters and that VP35 does not block signaling from the IFN-α/β receptor. The ability of VP35 to inhibit this virus-induced transcription correlates with its ability to block activation of IRF-3, a cellular transcription factor of central importance in initiating the host cell IFN response. We demonstrate that VP35 blocks the Sendai virus-induced activation of two promoters which can be directly activated by IRF-3, namely, the ISG54 promoter and the ISG56 promoter. Further, expression of VP35 prevents the IRF-3-dependent activation of the IFN-α4 promoter in response to viral infection. The inhibition of IRF-3 appears to occur through an inhibition of IRF-3 phosphorylation. VP35 blocks virus-induced IRF-3 phosphorylation and subsequent IRF-3 dimerization and nuclear translocation. Consistent with these observations, Ebola virus infection of Vero cells activated neither transcription from the ISG54 promoter nor nuclear accumulation of IRF-3. These data suggest that in Ebola virus-infected cells, VP35 inhibits the induction of antiviral genes, including the IFN-β gene, by blocking IRF-3 activation.

scholarly journals The IRF-3 Transcription Factor Mediates Sendai Virus-Induced Apoptosis

2000 ◽  
Vol 74 (8) ◽  
pp. 3781-3792 ◽  
Author(s):  
Christophe Heylbroeck ◽  
Siddharth Balachandran ◽  
Marc J. Servant ◽  
Carmela DeLuca ◽  
Glen N. Barber ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Virus Infection ◽  
Nuclear Translocation ◽  
Sendai Virus ◽  
Active Form ◽  
Cellular Transformation ◽  
Dominant Negative ◽  
Target Cells ◽  
Induced Apoptosis ◽  
Constitutively Active

ABSTRACT Virus infection of target cells can result in different biological outcomes: lytic infection, cellular transformation, or cell death by apoptosis. Cells respond to virus infection by the activation of specific transcription factors involved in cytokine gene regulation and cell growth control. The ubiquitously expressed interferon regulatory factor 3 (IRF-3) transcription factor is directly activated following virus infection through posttranslational modification. Phosphorylation of specific C-terminal serine residues results in IRF-3 dimerization, nuclear translocation, and activation of DNA-binding and transactivation potential. Once activated, IRF-3 transcriptionally up regulates alpha/beta interferon genes, the chemokine RANTES, and potentially other genes that inhibit viral infection. We previously generated constitutively active [IRF-3(5D)] and dominant negative (IRF-3 ΔN) forms of IRF-3 that control target gene expression. In an effort to characterize the growth regulatory properties of IRF-3, we observed that IRF-3 is a mediator of paramyxovirus-induced apoptosis. Expression of the constitutively active form of IRF-3 is toxic, preventing the establishment of stably transfected cells. By using a tetracycline-inducible system, we show that induction of IRF-3(5D) alone is sufficient to induce apoptosis in human embryonic kidney 293 and human Jurkat T cells as measured by DNA laddering, terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling assay, and analysis of DNA content by flow cytometry. Wild-type IRF-3 expression augments paramyxovirus-induced apoptosis, while expression of IRF-3 ΔN blocks virus-induced apoptosis. In addition, we demonstrate an important role of caspases 8, 9, and 3 in IRF-3-induced apoptosis. These results suggest that IRF-3, in addition to potently activating cytokine genes, regulates apoptotic signalling following virus infection.

scholarly journals PRV UL13 inhibits cGAS–STING-mediated IFN-β production by phosphorylating IRF3

2020 ◽  
Vol 51 (1) ◽  
Author(s):  
Zongyi Bo ◽  
Yurun Miao ◽  
Rui Xi ◽  
Qiuping Zhong ◽  
Chenyi Bao ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Pseudorabies Virus ◽  
Nuclear Translocation ◽  
Economic Loss ◽  
Inhibitory Effect ◽  
Poly I:C ◽  
Interferon Response ◽  
Type I ◽  
Creb Binding Protein ◽  
Swine Industry

Abstract Cyclic GMP-AMP (cGAMP) synthase (cGAS) is an intracellular sensor of cytoplasmic viral DNA created during virus infection, which subsequently activates the stimulator of interferon gene (STING)-dependent type I interferon response to eliminate pathogens. In contrast, viruses have developed different strategies to modulate this signalling pathway. Pseudorabies virus (PRV), an alphaherpesvirus, is the causative agent of Aujeszky’s disease (AD), a notable disease that causes substantial economic loss to the swine industry globally. Previous reports have shown that PRV infection induces cGAS-dependent IFN-β production, conversely hydrolysing cGAMP, a second messenger synthesized by cGAS, and attenuates PRV-induced IRF3 activation and IFN-β secretion. However, it is not clear whether PRV open reading frames (ORFs) modulate the cGAS–STING-IRF3 pathway. Here, 50 PRV ORFs were screened, showing that PRV UL13 serine/threonine kinase blocks the cGAS–STING-IRF3-, poly(I:C)- or VSV-mediated transcriptional activation of the IFN-β gene. Importantly, it was discovered that UL13 phosphorylates IRF3, and its kinase activity is indispensable for such an inhibitory effect. Moreover, UL13 does not affect IRF3 dimerization, nuclear translocation or association with CREB-binding protein (CBP) but attenuates the binding of IRF3 to the IRF3-responsive promoter. Consistent with this, it was discovered that UL13 inhibits the expression of multiple interferon-stimulated genes (ISGs) induced by cGAS–STING or poly(I:C). Finally, it was determined that PRV infection can activate IRF3 by recruiting it to the nucleus, and PRVΔUL13 mutants enhance the transactivation level of the IFN-β gene. Taken together, the data from the present study demonstrated that PRV UL13 inhibits cGAS–STING-mediated IFN-β production by phosphorylating IRF3.

scholarly journals Cubitus interruptus Requires DrosophilaCREB-Binding Protein To Activate wingless Expression in theDrosophila Embryo

2000 ◽  
Vol 20 (5) ◽  
pp. 1616-1625 ◽  
Author(s):  
Yang Chen ◽  
R. H. Goodman ◽  
Sarah M. Smolik
Keyword(s):  
Transcriptional Activation ◽  
Target Genes ◽  
Binding Protein ◽  
Point Mutations ◽  
Creb Binding Protein ◽  
Wild Type ◽  
Interaction Domain ◽  
Cubitus Interruptus

ABSTRACT CREB-binding protein (CBP) serves as a transcriptional coactivator in multiple signal transduction pathways. The Drosophilahomologue of CBP, dCBP, interacts with the transcription factors Cubitus interruptus (CI), MAD, and Dorsal (DL) and functions as a coactivator in several signaling pathways during Drosophiladevelopment, including the hedgehog (hh),decapentaplegic (dpp), and Tollpathways. Although dCBP is required for the expression of thehh target genes, wingless (wg) andpatched (ptc) in vivo, and potentiatesci-mediated transcriptional activation in vitro, it is not known that ci absolutely requires dCBP for its activity. We used a yeast genetic screen to identify several ci point mutations that disrupt CI-dCBP interactions. These mutant proteins are unable to transactivate a reporter gene regulated by cibinding sites and have a lower dCBP-stimulated activity than wild-type CI. When expressed exogenously in embryos, the CI point mutants cannot activate endogenous wg expression. Furthermore, a CI mutant protein that lacks the entire dCBP interaction domain functions as a negative competitor for wild-type CI activity, and the expression of dCBP antisense RNAs can suppress CI transactivation in Kc cells. Taken together, our data suggest that dCBP function is necessary forci-mediated transactivation of wg duringDrosophila embryogenesis.

scholarly journals Negative regulation of nuclear factor-kappaB activation and function by glucocorticoids

2002 ◽  
Vol 28 (2) ◽  
pp. 69-78 ◽  
Author(s):  
WY Almawi ◽  
OK Melemedjian
Keyword(s):  
Transcription Factors ◽  
Transcriptional Activation ◽  
Promoter Region ◽  
Nuclear Translocation ◽  
Nuclear Factor Kappab ◽  
Nuclear Factor ◽  
Creb Binding Protein ◽  
Transcriptional Initiation ◽  
Nf Kappab ◽  
And Function

Glucocorticoids (GCs) exert their anti-inflammatory and antiproliferative effects principally by inhibiting the expression of cytokines and adhesion molecules. Mechanistically, GCs diffuse through the cell membrane, and bind to their inactive cytosolic receptors (GRs), which then undergo conformational modifications that allow for their nuclear translocation. In the nucleus, activated GRs modulate transcriptional events by directly associating with DNA elements, compatible with the GCs response elements (GRE) motif, and located in variable copy numbers and at variable distances from the TATA box, in the promoter region of GC-responsive genes. In addition, activated GRs also acted by antagonizing the activity of transcription factors, in particular nuclear factor-kappaB (NF-kappaB), by direct and indirect mechanisms. GCs induced gene transcription and protein synthesis of the NF-kappaB inhibitor, IkappaB. Activated GR also antagonized NF-kappaB activity through protein-protein interaction involving direct complexing with, and inhibition of, NF-kappaB binding to DNA (Simple Model), or association with NF-kappaB bound to the kappaB DNA site (Composite Model). In addition, and according to the Transmodulation Model, GRE-bound GR may interact with and inhibit the activity of kappaB-bound NF-kappaB via a mechanism involving cross-talk between the two transcription factors. Lastly, GR may compete with NF-kappaB for nuclear coactivators, including CREB binding protein and p300, thereby reducing and inhibiting transcriptional activation by NF-kappaB. It should be noted that, in exerting its effect, activated GR did not affect the correct assembly of the pre-initiation (DAB) complex, but acted rather more proximally in inhibiting the correct assembly of transcription factors in the promoter region, and thus transcriptional initiation.

scholarly journals PCAF Interacts with Tax and Stimulates Tax Transactivation in a Histone Acetyltransferase-Independent Manner

1999 ◽  
Vol 19 (12) ◽  
pp. 8136-8145 ◽  
Author(s):  
Hua Jiang ◽  
Hanxin Lu ◽  
R. Louis Schiltz ◽  
Cynthia A. Pise-Masison ◽  
Vasily V. Ogryzko ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Histone Acetyltransferase ◽  
Point Mutations ◽  
Creb Binding Protein ◽  
Independent Manner ◽  
Cell Leukemia Virus Type ◽  
Knock Out ◽  
Human T Cell

ABSTRACT Recent studies have shown that the p300/CREB binding protein (CBP)-associated factor (PCAF) is involved in transcriptional activation. PCAF activity has been shown strongly associated with histone acetyltransferase (HAT) activity. In this report, we present evidence for a HAT-independent transcription function that is activated in the presence of the human T-cell leukemia virus type 1 (HTLV-1) Tax protein. In vitro and in vivo GST-Tax pull-down and coimmunoprecipitation experiments demonstrate that there is a direct interaction between Tax and PCAF, independent of p300/CBP. PCAF can be recruited to the HTLV-1 Tax responsive element in the presence of Tax, and PCAF cooperates with Tax in vivo to activate transcription from the HTLV-1 LTR over 10-fold. Point mutations at Tax amino acid 318 (TaxS318A) or 319 to 320 (Tax M47), which have decreased or no activity on the HTLV-1 promoter, are defective for PCAF binding. Strikingly, the ability of PCAF to stimulate Tax transactivation is not solely dependent on the PCAF HAT domain. Two independent PCAF HAT mutants, which knock out acetyltransferase enzyme activity, activate Tax transactivation to approximately the same level as wild-type PCAF. In contrast, p300 stimulation of Tax transactivation is HAT dependent. These studies provide experimental evidence that PCAF contains a coactivator transcription function independent of the HAT activity on the viral long terminal repeat.

scholarly journals A revised model for promoter competition based on multi-way chromatin interactions at the α-globin locus

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
A. Marieke Oudelaar ◽  
Caroline L. Harrold ◽  
Lars L. P. Hanssen ◽  
Jelena M. Telenius ◽  
Douglas R. Higgs ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Regulation Of Gene Expression ◽  
Regulatory Elements ◽  
Chromatin Domain ◽  
Gene Promoters ◽  
Multiple Promoters ◽  
Chromatin Interactions ◽  
Technological Advances ◽  
Single Complex ◽  
Membrane Bound

AbstractSpecific communication between gene promoters and enhancers is critical for accurate regulation of gene expression. However, it remains unclear how specific interactions between multiple regulatory elements contained within a single chromatin domain are coordinated. Recent technological advances which can detect multi-way chromatin interactions at single alleles can provide insights into how multiple regulatory elements cooperate or compete for transcriptional activation. Here, we use such an approach to investigate how interactions of the α-globin enhancers are distributed between multiple promoters in a mouse model in which the α-globin domain is extended to include several additional genes. Our data show that gene promoters do not form mutually exclusive interactions with enhancers, but all interact simultaneously in a single complex. These findings suggest that promoters do not structurally compete for interactions with enhancers, but form a regulatory hub structure, which is consistent with recent models of transcriptional activation occurring in non-membrane bound nuclear compartments.

scholarly journals The transcription factor GLI1 cooperates with the chromatin remodeler SMARCA2 to regulate chromatin accessibility at distal DNA regulatory elements

2020 ◽  
Vol 295 (26) ◽  
pp. 8725-8735
Author(s):  
Stephanie L. Safgren ◽  
Rachel L. O. Olson ◽  
Anne M. Vrabel ◽  
Luciana L. Almada ◽  
David L. Marks ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Transcriptional Activation ◽  
Transcriptional Activity ◽  
Regulatory Elements ◽  
Chromatin Accessibility ◽  
Micrococcal Nuclease ◽  
Gene Promoters ◽  
Interacting Protein ◽  
Transcriptional Activation Domain ◽  
Dna Regulatory Elements

The transcription factor GLI1 (GLI family zinc finger 1) plays a key role in the development and progression of multiple malignancies. To date, regulation of transcriptional activity at target gene promoters is the only molecular event known to underlie the oncogenic function of GLI1. Here, we provide evidence that GLI1 controls chromatin accessibility at distal regulatory regions by modulating the recruitment of SMARCA2 (SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily A, member 2) to these elements. We demonstrate that SMARCA2 endogenously interacts with GLI1 and enhances its transcriptional activity. Mapping experiments indicated that the C-terminal transcriptional activation domain of GLI1 and SMARCA2's central domains, including its ATPase motif, are required for this interaction. Interestingly, similar to SMARCA2, GLI1 overexpression increased chromatin accessibility, as indicated by results of the micrococcal nuclease assay. Further, results of assays for transposase-accessible chromatin with sequencing (ATAC-seq) after GLI1 knockdown supported these findings, revealing that GLI1 regulates chromatin accessibility at several regions distal to gene promoters. Integrated RNA-seq and ATAC-seq data analyses identified a subset of differentially expressed genes located in cis to these regulated chromatin sites. Finally, using the GLI1-regulated gene HHIP (Hedgehog-interacting protein) as a model, we demonstrate that GLI1 and SMARCA2 co-occupy a distal chromatin peak and that SMARCA2 recruitment to this HHIP putative enhancer requires intact GLI1. These findings provide insights into how GLI1 controls gene expression in cancer cells and may inform approaches targeting this oncogenic transcription factor to manage malignancies.

scholarly journals Myeloid neddylation targets IRF7 and promotes host innate immunity against RNA viruses

2021 ◽  
Vol 17 (9) ◽  
pp. e1009901
Author(s):  
Min Zhao ◽  
Yaolin Zhang ◽  
Xiqin Yang ◽  
Jiayang Jin ◽  
Zhuo Shen ◽  
...  
Keyword(s):  
Innate Immunity ◽  
Virus Infection ◽  
Nuclear Translocation ◽  
Rna Viruses ◽  
Rna Virus ◽  
Innate Immune ◽  
Type I ◽  
Gene Promoters ◽  
Post Translational Modification

Neddylation, an important type of post-translational modification, has been implicated in innate and adapted immunity. But the role of neddylation in innate immune response against RNA viruses remains elusive. Here we report that neddylation promotes RNA virus-induced type I IFN production, especially IFN-α. More importantly, myeloid deficiency of UBA3 or NEDD8 renders mice less resistant to RNA virus infection. Neddylation is essential for RNA virus-triggered activation of Ifna gene promoters. Further exploration has revealed that mammalian IRF7undergoes neddylation, which is enhanced after RNA virus infection. Even though neddylation blockade does not hinder RNA virus-triggered IRF7 expression, IRF7 mutant defective in neddylation exhibits reduced ability to activate Ifna gene promoters. Neddylation blockade impedes RNA virus-induced IRF7 nuclear translocation without hindering its phosphorylation and dimerization with IRF3. By contrast, IRF7 mutant defective in neddylation shows enhanced dimerization with IRF5, an Ifna repressor when interacting with IRF7. In conclusion, our data demonstrate that myeloid neddylation contributes to host anti-viral innate immunity through targeting IRF7 and promoting its transcriptional activity.

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