Extensive Cooperation of Immune Master Regulators IRF3 and NFκB in RNA Pol II Recruitment and Pause Release in Human Innate Antiviral Transcription

Freaney, Jonathan E.; Kim, Rebecca; Mandhana, Roli; Horvath, Curt M.

doi:10.1016/j.celrep.2013.07.043

Cited by 69 publications

(75 citation statements)

References 65 publications

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“…18,19,31,32,43 Conversely, RelA, as well as other NF- κ B subunits, can modulate the DNA binding properties of other transcription factors. 42,43 Thus, in addition to their well-studied direct DNA binding properties, each of these transcription factors appears to function in the nucleus as a signaling cofactor that influences the DNA binding and transcriptional potential of their binding partners even at locations within the genome where no consensus DNA binding site for the cofactor is present.…”

Section: Discussionmentioning

confidence: 99%

Protein Cofactors Are Essential for High-Affinity DNA Binding by the Nuclear Factor κB RelA Subunit

Mulero

Shahabi

et al. 2018

Biochemistry

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Transcription activator proteins typically contain two functional domains: a DNA binding domain (DBD) that binds to DNA with sequence specificity and an activation domain (AD) whose established function is to recruit RNA polymerase. In this report, we show that purified recombinant nuclear factor κB (NF-κB) RelA dimers bind specific κB DNA sites with an affinity significantly lower than that of the same dimers from nuclear extracts of activated cells, suggesting that additional nuclear cofactors might facilitate DNA binding by the RelA dimers. Additionally, recombinant RelA binds DNA with relatively low affinity at a physiological salt concentration in vitro. The addition of p53 or RPS3 (ribosomal protein S3) increases RelA:DNA binding affinity 2- to >50-fold depending on the protein and ionic conditions. These cofactor proteins do not form stable ternary complexes, suggesting that they stabilize the RelA:DNA complex through dynamic interactions. Surprisingly, the RelA-DBD alone fails to bind DNA under the same solution conditions even in the presence of cofactors, suggesting an important role of the RelA-AD in DNA binding. Reduced RelA:DNA binding at a physiological ionic strength suggests that multiple cofactors might be acting simultaneously to mitigate the electrolyte effect and stabilize the RelA:DNA complex in vivo. Overall, our observations suggest that the RelA-AD and multiple cofactor proteins function cooperatively to prime the RelA-DBD and stabilize the RelA:DNA complex in cells. Our study provides a mechanism for nuclear cofactor proteins in NF-κB-dependent gene regulation.

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Section: Discussionmentioning

confidence: 99%

Protein Cofactors Are Essential for High-Affinity DNA Binding by the Nuclear Factor κB RelA Subunit

Mulero

Shahabi

et al. 2018

Biochemistry

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“…Based on these findings, and given the fact that many developmental and immune genes are regulated by transcriptional pausing (4,6,8,14,23,24), we hypothesized that Nup98 also may regulate virus-induced antiviral genes. We found that Nup98 plays a broadly antiviral role in insects; cells or adult flies depleted of Nup98 are more susceptible to infection against a panel of disparate RNA viruses.…”

Section: Significancementioning

confidence: 99%

Nup98 promotes antiviral gene expression to restrict RNA viral infection in Drosophila

Panda

Pascual-Garcia²,

Dunagin

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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In response to infection, the innate immune system rapidly activates an elaborate and tightly orchestrated gene expression program to induce critical antimicrobial genes. While many key players in this program have been identified in disparate biological systems, it is clear that there are additional uncharacterized mechanisms at play. Our previous studies revealed that a rapidly-induced antiviral gene expression program is active against disparate human arthropod-borne viruses in Drosophila. Moreover, one-half of this program is regulated at the level of transcriptional pausing. Here we found that Nup98, a virus-induced gene, was antiviral against a panel of viruses both in cells and adult flies since its depletion significantly enhanced viral infection. Mechanistically, we found that Nup98 promotes antiviral gene expression in Drosophila at the level of transcription. Expression profiling revealed that the virus-induced activation of 36 genes was abrogated upon loss of Nup98; and we found that a subset of these Nup98-dependent genes were antiviral. These Nup98-dependent virus-induced genes are Cdk9-dependent and translation-independent suggesting that these are rapidly induced primary response genes. Biochemically, we demonstrate that Nup98 is directly bound to the promoters of virus-induced genes, and that it promotes occupancy of the initiating form of RNA polymerase II at these promoters, which are rapidly induced on viral infection to restrict human arboviruses in insects.innate immunity | nuclear pore | nucleoporin | pol II regulation | P-TEFb

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“…We propose that the cGAS/STING/TBK1 cascade is an important contributor to basal maintenance of a primed state for ISGs. Interferon sensitization has been hypothesized to contribute to maintenance of a basal state priming for responsive cell lines that facilitates rapid induction of target genes (46)(47)(48). Changes in basal priming of ISGs may contribute to alterations in the antiviral response phenotype and provide an avenue for modifying the antiviral response in a cell-specific manner.…”

Section: Figmentioning

confidence: 99%

Adenovirus Detection by the cGAS/STING/TBK1 DNA Sensing Cascade

Lam

Stein

Falck-Pedersen

2014

J Virol

200

149

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T he human serotype 5 adenovirus (Ad5) is a nonenveloped linear double-stranded DNA virus associated with upper respiratory tract disease in humans. It has been extensively studied as a model for virus and host cell interactions. Replication-defective recombinant Ad5 vectors (rAdV) deleted in E1 and E3 coding domains have been characterized in gene therapy, vaccine, and oncolytic vector strategies in the murine model. Although nonpermissive for Ad5 replication, the murine model of rAdV infection provides a valuable resource for characterizing how the innate and adaptive immune response orchestrates an antiviral response to nonenveloped DNA viruses.Virus uptake by immune sentinel cells such as macrophage and dendritic cells is vital to initiating the antiviral immune response. In addition to antigen-presenting cells (APCs), other cell types, including endothelial cells or tissue-specific cells such as hepatocytes, when exposed to virus, also contribute to the host antiviral response. In vitro studies of isolated bone marrow-derived APCs or representative cell lines have revealed a cell-specific antiviral innate response, where activation of the type I interferon (IFN) cascade is a dominant feature (1-4). A valuable marker for early events in the antiviral recognition response is activation of the transcription factor interferon response factor 3 (IRF3). Following infection, cytosolic IRF3 undergoes phosphorylation as a primary response to adenovirus uptake. Activation occurs in a MyD88/TRIF-independent manner; it requires integrin-dependent endosomal entry, escape, and presentation of viral DNA to the cytosolic compartment (3).In rAdV-responsive murine cell lines, the STING/TBK1 cascade is required for IRF3 phosphorylation (5, 6). STING (7,8) functions as an adaptor linking DNA recognition signaling to activation of the TBK1 kinase. TBK1 activation (9) leads to C-terminal IRF3 phosphorylation, dimerization, and translocation to the nucleus (10, 11). In the nucleus, IRF3, in collaboration with additional transcription factors (NF-B and AP1), results in transcriptional activation of IRF3-responsive genes (including IFN-␤) (12). This sequence of events contributes to the primary antiviral response to adenovirus infection. The translation of primary response transcripts such as IFN-␤ leads to autocrine/paracrine secondary signaling. The combination of primary and secondary response functions leads to expression of a complete antiviral response, which is distinct for different cell types.Using various screening protocols, cell lines, and output assays, an extensive list of cytosolic DNA sensors, including DAI, RNA polymerase (Pol) III, IFI16, DDX41, and Aim 2, has been established (reviewed in reference 13). However, the DNA sensor involved in recognizing infection by adenovirus leading to early IRF3 activation has not been convincingly established. The recent identification of cyclic dinucleotide activation of STING (14-18) and the elegant discovery of cyclic-GMP-AMP synthase (cGAS) as a DNA sensor (19,20) provide an...

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Extensive Cooperation of Immune Master Regulators IRF3 and NFκB in RNA Pol II Recruitment and Pause Release in Human Innate Antiviral Transcription

Cited by 69 publications

References 65 publications

Protein Cofactors Are Essential for High-Affinity DNA Binding by the Nuclear Factor κB RelA Subunit

Protein Cofactors Are Essential for High-Affinity DNA Binding by the Nuclear Factor κB RelA Subunit

Nup98 promotes antiviral gene expression to restrict RNA viral infection in Drosophila

Adenovirus Detection by the cGAS/STING/TBK1 DNA Sensing Cascade

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