Protein kinase R mediates intestinal epithelial gene remodeling in response to double-stranded RNA and live rotavirus

Matam Vijay-Kumar, Jon R. Gentsch, William Kaiser, Niels Borregaard, Margaret K. Offermann, Andrew S. Neish, Andrew T. Gewirtz

Research output: Contribution to journalArticle

37 Citations (Scopus)

Abstract

As sentinels of host defense, intestinal epithelial cells respond to the viral pathogen rotavirus by activating a gene expression that promotes immune cell recruitment and activation. We hypothesized that epithelial sensing of rotavirus might target dsRNA, which can be detected by TLR3 or protein kinase R (PKR). Accordingly, we observed that synthetic dsRNA, polyinosinic acid:cytidylic acid (poly(I:C)), potently induced gene remodeling in model intestinal epithelia with the specific pattern of expressed genes, including both classic proinflammatory genes (e.g., IL-8), as well as genes that are classically activated in virus-infected cells (e.g., IFN-responsive genes). Poly(I:C)-induced IL-8 was concentration dependent (2-100 μg/ml) and displayed slower kinetics compared with IL-8 induced by bacterial flagellin (ET50 ∼24 vs 8 h poly(I:C) vs flagellin, respectively). Although model epithelia expressed detectable TLR3 mRNA, neither TLR3-neutralizing Abs nor chloroquine, which blocks activation of intracellular TLR3, attenuated epithelial responses to poly(I:C). Conversely, poly(I:C)-induced phosphorylation of PKR and inhibitors of PKR, 2-aminopurine and adenine, ablated poly(I:C)-induced gene expression but had no effect on gene expression induced by flagellin, thus suggesting that intestinal epithelial cell detection of dsRNA relies on PKR. Consistent with poly(I:C) detection by an intracellular molecule such as PKR, we observed that both uptake of and responses to poly(I:C) were polarized to the basolateral side. Lastly, we observed that the pattern of pharmacologic inhibition of responses to poly(I:C) was identical to that seen in response to infection by live rotavirus, indicating a potentially important role for PKR in activating intestinal epithelial gene expression in rotavirus infection.

Original languageEnglish (US)
Pages (from-to)6322-6331
Number of pages10
JournalJournal of Immunology
Volume174
Issue number10
DOIs
StatePublished - May 15 2005
Externally publishedYes

Fingerprint

Poly I
Cytidine Monophosphate
Double-Stranded RNA
Rotavirus
Protein Kinases
Flagellin
Genes
Interleukin-8
Rotavirus Infections
Gene Expression
2-Aminopurine
Epithelial Cells
Chloroquine
Adenine
Intestinal Mucosa
Protein Kinase Inhibitors
Epithelium
Phosphorylation
Viruses
Messenger RNA

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

Cite this

Vijay-Kumar, M., Gentsch, J. R., Kaiser, W., Borregaard, N., Offermann, M. K., Neish, A. S., & Gewirtz, A. T. (2005). Protein kinase R mediates intestinal epithelial gene remodeling in response to double-stranded RNA and live rotavirus. Journal of Immunology, 174(10), 6322-6331. https://doi.org/10.4049/jimmunol.174.10.6322

Protein kinase R mediates intestinal epithelial gene remodeling in response to double-stranded RNA and live rotavirus. / Vijay-Kumar, Matam; Gentsch, Jon R.; Kaiser, William; Borregaard, Niels; Offermann, Margaret K.; Neish, Andrew S.; Gewirtz, Andrew T.

In: Journal of Immunology, Vol. 174, No. 10, 15.05.2005, p. 6322-6331.

Research output: Contribution to journalArticle

Vijay-Kumar, Matam ; Gentsch, Jon R. ; Kaiser, William ; Borregaard, Niels ; Offermann, Margaret K. ; Neish, Andrew S. ; Gewirtz, Andrew T. / Protein kinase R mediates intestinal epithelial gene remodeling in response to double-stranded RNA and live rotavirus. In: Journal of Immunology. 2005 ; Vol. 174, No. 10. pp. 6322-6331.
@article{2a2739f2bcc34637851ac0bfd56b88b4,
title = "Protein kinase R mediates intestinal epithelial gene remodeling in response to double-stranded RNA and live rotavirus",
abstract = "As sentinels of host defense, intestinal epithelial cells respond to the viral pathogen rotavirus by activating a gene expression that promotes immune cell recruitment and activation. We hypothesized that epithelial sensing of rotavirus might target dsRNA, which can be detected by TLR3 or protein kinase R (PKR). Accordingly, we observed that synthetic dsRNA, polyinosinic acid:cytidylic acid (poly(I:C)), potently induced gene remodeling in model intestinal epithelia with the specific pattern of expressed genes, including both classic proinflammatory genes (e.g., IL-8), as well as genes that are classically activated in virus-infected cells (e.g., IFN-responsive genes). Poly(I:C)-induced IL-8 was concentration dependent (2-100 μg/ml) and displayed slower kinetics compared with IL-8 induced by bacterial flagellin (ET50 ∼24 vs 8 h poly(I:C) vs flagellin, respectively). Although model epithelia expressed detectable TLR3 mRNA, neither TLR3-neutralizing Abs nor chloroquine, which blocks activation of intracellular TLR3, attenuated epithelial responses to poly(I:C). Conversely, poly(I:C)-induced phosphorylation of PKR and inhibitors of PKR, 2-aminopurine and adenine, ablated poly(I:C)-induced gene expression but had no effect on gene expression induced by flagellin, thus suggesting that intestinal epithelial cell detection of dsRNA relies on PKR. Consistent with poly(I:C) detection by an intracellular molecule such as PKR, we observed that both uptake of and responses to poly(I:C) were polarized to the basolateral side. Lastly, we observed that the pattern of pharmacologic inhibition of responses to poly(I:C) was identical to that seen in response to infection by live rotavirus, indicating a potentially important role for PKR in activating intestinal epithelial gene expression in rotavirus infection.",
author = "Matam Vijay-Kumar and Gentsch, {Jon R.} and William Kaiser and Niels Borregaard and Offermann, {Margaret K.} and Neish, {Andrew S.} and Gewirtz, {Andrew T.}",
year = "2005",
month = "5",
day = "15",
doi = "10.4049/jimmunol.174.10.6322",
language = "English (US)",
volume = "174",
pages = "6322--6331",
journal = "Journal of Immunology",
issn = "0022-1767",
publisher = "American Association of Immunologists",
number = "10",

}

TY - JOUR

T1 - Protein kinase R mediates intestinal epithelial gene remodeling in response to double-stranded RNA and live rotavirus

AU - Vijay-Kumar, Matam

AU - Gentsch, Jon R.

AU - Kaiser, William

AU - Borregaard, Niels

AU - Offermann, Margaret K.

AU - Neish, Andrew S.

AU - Gewirtz, Andrew T.

PY - 2005/5/15

Y1 - 2005/5/15

N2 - As sentinels of host defense, intestinal epithelial cells respond to the viral pathogen rotavirus by activating a gene expression that promotes immune cell recruitment and activation. We hypothesized that epithelial sensing of rotavirus might target dsRNA, which can be detected by TLR3 or protein kinase R (PKR). Accordingly, we observed that synthetic dsRNA, polyinosinic acid:cytidylic acid (poly(I:C)), potently induced gene remodeling in model intestinal epithelia with the specific pattern of expressed genes, including both classic proinflammatory genes (e.g., IL-8), as well as genes that are classically activated in virus-infected cells (e.g., IFN-responsive genes). Poly(I:C)-induced IL-8 was concentration dependent (2-100 μg/ml) and displayed slower kinetics compared with IL-8 induced by bacterial flagellin (ET50 ∼24 vs 8 h poly(I:C) vs flagellin, respectively). Although model epithelia expressed detectable TLR3 mRNA, neither TLR3-neutralizing Abs nor chloroquine, which blocks activation of intracellular TLR3, attenuated epithelial responses to poly(I:C). Conversely, poly(I:C)-induced phosphorylation of PKR and inhibitors of PKR, 2-aminopurine and adenine, ablated poly(I:C)-induced gene expression but had no effect on gene expression induced by flagellin, thus suggesting that intestinal epithelial cell detection of dsRNA relies on PKR. Consistent with poly(I:C) detection by an intracellular molecule such as PKR, we observed that both uptake of and responses to poly(I:C) were polarized to the basolateral side. Lastly, we observed that the pattern of pharmacologic inhibition of responses to poly(I:C) was identical to that seen in response to infection by live rotavirus, indicating a potentially important role for PKR in activating intestinal epithelial gene expression in rotavirus infection.

AB - As sentinels of host defense, intestinal epithelial cells respond to the viral pathogen rotavirus by activating a gene expression that promotes immune cell recruitment and activation. We hypothesized that epithelial sensing of rotavirus might target dsRNA, which can be detected by TLR3 or protein kinase R (PKR). Accordingly, we observed that synthetic dsRNA, polyinosinic acid:cytidylic acid (poly(I:C)), potently induced gene remodeling in model intestinal epithelia with the specific pattern of expressed genes, including both classic proinflammatory genes (e.g., IL-8), as well as genes that are classically activated in virus-infected cells (e.g., IFN-responsive genes). Poly(I:C)-induced IL-8 was concentration dependent (2-100 μg/ml) and displayed slower kinetics compared with IL-8 induced by bacterial flagellin (ET50 ∼24 vs 8 h poly(I:C) vs flagellin, respectively). Although model epithelia expressed detectable TLR3 mRNA, neither TLR3-neutralizing Abs nor chloroquine, which blocks activation of intracellular TLR3, attenuated epithelial responses to poly(I:C). Conversely, poly(I:C)-induced phosphorylation of PKR and inhibitors of PKR, 2-aminopurine and adenine, ablated poly(I:C)-induced gene expression but had no effect on gene expression induced by flagellin, thus suggesting that intestinal epithelial cell detection of dsRNA relies on PKR. Consistent with poly(I:C) detection by an intracellular molecule such as PKR, we observed that both uptake of and responses to poly(I:C) were polarized to the basolateral side. Lastly, we observed that the pattern of pharmacologic inhibition of responses to poly(I:C) was identical to that seen in response to infection by live rotavirus, indicating a potentially important role for PKR in activating intestinal epithelial gene expression in rotavirus infection.

UR - http://www.scopus.com/inward/record.url?scp=18644365496&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=18644365496&partnerID=8YFLogxK

U2 - 10.4049/jimmunol.174.10.6322

DO - 10.4049/jimmunol.174.10.6322

M3 - Article

C2 - 15879132

AN - SCOPUS:18644365496

VL - 174

SP - 6322

EP - 6331

JO - Journal of Immunology

JF - Journal of Immunology

SN - 0022-1767

IS - 10

ER -