CDK9-dependent transcriptional elongation in the innate interferon-stimulated gene response to respiratory syncytial virus infection in airway epithelial cells

Bing Tian, Yingxin Zhao, Mridul Kalita, Chukwudi B. Edeh, Slobodan Paessler, Antonella Casola, Michael N. Teng, Roberto Garofalo, Allan R. Brasier

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Abstract

Respiratory syncytial virus (RSV) is a negative-sense single-stranded RNA virus responsible for lower respiratory tract infections. During infection, the presence of double-stranded RNA (dsRNA) activates the interferon (IFN) regulatory factor 3 (IRF3) transcription factor, an event triggering expression of immediate early, IFN-stimulated genes (ISGs). We examine the role of transcriptional elongation in control of IRF3-dependent ISG expression. RSV infection induces ISG54, ISG56, and CIG5 gene expression in an IRF3-dependent manner demonstrated by IRF3 small interfering RNA (siRNA) silencing in both A549 epithelial cells and IRF3-/- MEFs. ISG expression was mediated by the recruitment of IRF3, CDK9, polymerase II (Pol II), and phospho- Ser2 carboxy-terminal domain (CTD) Pol II to the IFN-stimulated response element (ISRE) binding sites of the IRF3-dependent ISG promoters in native chromatin. We find that RSV infection enhances the activated fraction of cyclin-dependent kinase 9 (CDK9) by promoting its association with bromodomain 4 (BRD4) and disrupting its association with the inhibitory 7SK small nuclear RNA. The requirement of CDK9 activity for ISG expression was shown by siRNA-mediated silencing of CDK9 and by a selective CDK9 inhibitor in A549 cells. In contrast, RSV-induced beta interferon (IFN-ß) expression is not influenced by CDK9 inhibition. Using transcript-selective quantitative real-time reverse transcription-PCR (Q-RT-PCR) assays for the ISG54 gene, we observed that RSV induces transition from short to fully spliced mRNA transcripts and that this transition is blocked by CDK9 inhibition in both A549 and primary human small airway epithelial cells. These data indicate that transcription elongation plays a major role in RSV-induced ISG expression and is mediated by IRF3-dependent recruitment of activated CDK9. CDK9 activity may be a target for immunomodulation in RSV-induced lung disease.

Original languageEnglish (US)
Pages (from-to)7075-7092
Number of pages18
JournalJournal of Virology
Volume87
Issue number12
DOIs
StatePublished - Jun 2013

Fingerprint

Cyclin-Dependent Kinase 9
Interferon Regulatory Factor-3
Respiratory Syncytial Virus Infections
cyclin-dependent kinase
interferons
Interferons
epithelial cells
Epithelial Cells
Respiratory Syncytial Viruses
viruses
infection
Genes
genes
Gene Expression
gene expression
small interfering RNA
RNA Interference
respiratory tract diseases
Small Interfering RNA
negative sense, single-stranded RNA viruses

ASJC Scopus subject areas

  • Immunology
  • Virology

Cite this

@article{179b19e54f1c4c348efabb19e5f4f2dd,
title = "CDK9-dependent transcriptional elongation in the innate interferon-stimulated gene response to respiratory syncytial virus infection in airway epithelial cells",
abstract = "Respiratory syncytial virus (RSV) is a negative-sense single-stranded RNA virus responsible for lower respiratory tract infections. During infection, the presence of double-stranded RNA (dsRNA) activates the interferon (IFN) regulatory factor 3 (IRF3) transcription factor, an event triggering expression of immediate early, IFN-stimulated genes (ISGs). We examine the role of transcriptional elongation in control of IRF3-dependent ISG expression. RSV infection induces ISG54, ISG56, and CIG5 gene expression in an IRF3-dependent manner demonstrated by IRF3 small interfering RNA (siRNA) silencing in both A549 epithelial cells and IRF3-/- MEFs. ISG expression was mediated by the recruitment of IRF3, CDK9, polymerase II (Pol II), and phospho- Ser2 carboxy-terminal domain (CTD) Pol II to the IFN-stimulated response element (ISRE) binding sites of the IRF3-dependent ISG promoters in native chromatin. We find that RSV infection enhances the activated fraction of cyclin-dependent kinase 9 (CDK9) by promoting its association with bromodomain 4 (BRD4) and disrupting its association with the inhibitory 7SK small nuclear RNA. The requirement of CDK9 activity for ISG expression was shown by siRNA-mediated silencing of CDK9 and by a selective CDK9 inhibitor in A549 cells. In contrast, RSV-induced beta interferon (IFN-{\ss}) expression is not influenced by CDK9 inhibition. Using transcript-selective quantitative real-time reverse transcription-PCR (Q-RT-PCR) assays for the ISG54 gene, we observed that RSV induces transition from short to fully spliced mRNA transcripts and that this transition is blocked by CDK9 inhibition in both A549 and primary human small airway epithelial cells. These data indicate that transcription elongation plays a major role in RSV-induced ISG expression and is mediated by IRF3-dependent recruitment of activated CDK9. CDK9 activity may be a target for immunomodulation in RSV-induced lung disease.",
author = "Bing Tian and Yingxin Zhao and Mridul Kalita and Edeh, {Chukwudi B.} and Slobodan Paessler and Antonella Casola and Teng, {Michael N.} and Roberto Garofalo and Brasier, {Allan R.}",
year = "2013",
month = "6",
doi = "10.1128/JVI.03399-12",
language = "English (US)",
volume = "87",
pages = "7075--7092",
journal = "Journal of Virology",
issn = "0022-538X",
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TY - JOUR

T1 - CDK9-dependent transcriptional elongation in the innate interferon-stimulated gene response to respiratory syncytial virus infection in airway epithelial cells

AU - Tian, Bing

AU - Zhao, Yingxin

AU - Kalita, Mridul

AU - Edeh, Chukwudi B.

AU - Paessler, Slobodan

AU - Casola, Antonella

AU - Teng, Michael N.

AU - Garofalo, Roberto

AU - Brasier, Allan R.

PY - 2013/6

Y1 - 2013/6

N2 - Respiratory syncytial virus (RSV) is a negative-sense single-stranded RNA virus responsible for lower respiratory tract infections. During infection, the presence of double-stranded RNA (dsRNA) activates the interferon (IFN) regulatory factor 3 (IRF3) transcription factor, an event triggering expression of immediate early, IFN-stimulated genes (ISGs). We examine the role of transcriptional elongation in control of IRF3-dependent ISG expression. RSV infection induces ISG54, ISG56, and CIG5 gene expression in an IRF3-dependent manner demonstrated by IRF3 small interfering RNA (siRNA) silencing in both A549 epithelial cells and IRF3-/- MEFs. ISG expression was mediated by the recruitment of IRF3, CDK9, polymerase II (Pol II), and phospho- Ser2 carboxy-terminal domain (CTD) Pol II to the IFN-stimulated response element (ISRE) binding sites of the IRF3-dependent ISG promoters in native chromatin. We find that RSV infection enhances the activated fraction of cyclin-dependent kinase 9 (CDK9) by promoting its association with bromodomain 4 (BRD4) and disrupting its association with the inhibitory 7SK small nuclear RNA. The requirement of CDK9 activity for ISG expression was shown by siRNA-mediated silencing of CDK9 and by a selective CDK9 inhibitor in A549 cells. In contrast, RSV-induced beta interferon (IFN-ß) expression is not influenced by CDK9 inhibition. Using transcript-selective quantitative real-time reverse transcription-PCR (Q-RT-PCR) assays for the ISG54 gene, we observed that RSV induces transition from short to fully spliced mRNA transcripts and that this transition is blocked by CDK9 inhibition in both A549 and primary human small airway epithelial cells. These data indicate that transcription elongation plays a major role in RSV-induced ISG expression and is mediated by IRF3-dependent recruitment of activated CDK9. CDK9 activity may be a target for immunomodulation in RSV-induced lung disease.

AB - Respiratory syncytial virus (RSV) is a negative-sense single-stranded RNA virus responsible for lower respiratory tract infections. During infection, the presence of double-stranded RNA (dsRNA) activates the interferon (IFN) regulatory factor 3 (IRF3) transcription factor, an event triggering expression of immediate early, IFN-stimulated genes (ISGs). We examine the role of transcriptional elongation in control of IRF3-dependent ISG expression. RSV infection induces ISG54, ISG56, and CIG5 gene expression in an IRF3-dependent manner demonstrated by IRF3 small interfering RNA (siRNA) silencing in both A549 epithelial cells and IRF3-/- MEFs. ISG expression was mediated by the recruitment of IRF3, CDK9, polymerase II (Pol II), and phospho- Ser2 carboxy-terminal domain (CTD) Pol II to the IFN-stimulated response element (ISRE) binding sites of the IRF3-dependent ISG promoters in native chromatin. We find that RSV infection enhances the activated fraction of cyclin-dependent kinase 9 (CDK9) by promoting its association with bromodomain 4 (BRD4) and disrupting its association with the inhibitory 7SK small nuclear RNA. The requirement of CDK9 activity for ISG expression was shown by siRNA-mediated silencing of CDK9 and by a selective CDK9 inhibitor in A549 cells. In contrast, RSV-induced beta interferon (IFN-ß) expression is not influenced by CDK9 inhibition. Using transcript-selective quantitative real-time reverse transcription-PCR (Q-RT-PCR) assays for the ISG54 gene, we observed that RSV induces transition from short to fully spliced mRNA transcripts and that this transition is blocked by CDK9 inhibition in both A549 and primary human small airway epithelial cells. These data indicate that transcription elongation plays a major role in RSV-induced ISG expression and is mediated by IRF3-dependent recruitment of activated CDK9. CDK9 activity may be a target for immunomodulation in RSV-induced lung disease.

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