Activation of RAW264.7 Macrophages by Bacterial DNA and Lipopolysaccharide Increases Cell Surface DNA Binding and Internalization

Sharon L. McCoy, Stephen E. Kurtz, Frances A. Hausman, Dennis Trune, Robert (Rob) Bennett, Steven Hefeneider

Research output: Contribution to journalArticle

32 Citations (Scopus)

Abstract

Bacterial DNA containing unmethylated CpG motifs is a pathogen-associated molecular pattern (PAMP) that interacts with host immune cells via a toll-like receptor (TLR) to induce immune responses. DNA binding and internalization into cells is independent of TLR expression, receptor-mediated, and required for cell activation. The objective of this study was to determine whether exposure of immune cells to bacterial DNA affects DNA binding and internalization. Treatment of RAW264.7 cells with CpG oligodeoxynucleotide (ODN) for both 18 and 42 h resulted in a significant increase in DNA binding, whereas non-CpG ODN had no effect on DNA binding. Enhanced DNA binding was non-sequence-specific, inhibited by unlabeled DNA, showed saturation, was consistent with increased cell surface DNA receptors, and resulted in enhanced internalization of DNA. Treatment with Escherichia coli DNA or lipopolysaccharide (LPS) also resulted in a significant increase in DNA binding, but treatment with interleukin-1α, tumor necrosis factor-α, or phorbol 12-myristate 13-acetate had no effect on DNA binding. Soluble factors produced in response to treatment with CpG ODN or LPS did not affect DNA binding. These studies demonstrate that one consequence of activating the host innate immune response by bacterial infection is enhanced binding and internalization of DNA. During this period of increased DNA internalization, RAW264.7 cells were hypo-responsive to continued stimulation by CpG ODN, as assessed by tumor necrosis factor-α activity. We speculate the biological significance of increasing DNA binding and internalization following interaction with bacterial PAMPs may provide a mechanism to limit an ongoing immune inflammatory response by enhancing clearance of bacterial DNA from the extracellular environment.

Original languageEnglish (US)
Pages (from-to)17217-17223
Number of pages7
JournalJournal of Biological Chemistry
Volume279
Issue number17
DOIs
StatePublished - Apr 23 2004

Fingerprint

Bacterial DNA
Macrophage Activation
Macrophages
Lipopolysaccharides
Chemical activation
DNA
Oligodeoxyribonucleotides
Toll-Like Receptors
Tumor Necrosis Factor-alpha
Cell Surface Receptors
Interleukin-1
Bacterial Infections
Innate Immunity
Escherichia coli

ASJC Scopus subject areas

  • Biochemistry

Cite this

Activation of RAW264.7 Macrophages by Bacterial DNA and Lipopolysaccharide Increases Cell Surface DNA Binding and Internalization. / McCoy, Sharon L.; Kurtz, Stephen E.; Hausman, Frances A.; Trune, Dennis; Bennett, Robert (Rob); Hefeneider, Steven.

In: Journal of Biological Chemistry, Vol. 279, No. 17, 23.04.2004, p. 17217-17223.

Research output: Contribution to journalArticle

McCoy, Sharon L. ; Kurtz, Stephen E. ; Hausman, Frances A. ; Trune, Dennis ; Bennett, Robert (Rob) ; Hefeneider, Steven. / Activation of RAW264.7 Macrophages by Bacterial DNA and Lipopolysaccharide Increases Cell Surface DNA Binding and Internalization. In: Journal of Biological Chemistry. 2004 ; Vol. 279, No. 17. pp. 17217-17223.
@article{a9ba1f3824124f76a87ee2f233dad623,
title = "Activation of RAW264.7 Macrophages by Bacterial DNA and Lipopolysaccharide Increases Cell Surface DNA Binding and Internalization",
abstract = "Bacterial DNA containing unmethylated CpG motifs is a pathogen-associated molecular pattern (PAMP) that interacts with host immune cells via a toll-like receptor (TLR) to induce immune responses. DNA binding and internalization into cells is independent of TLR expression, receptor-mediated, and required for cell activation. The objective of this study was to determine whether exposure of immune cells to bacterial DNA affects DNA binding and internalization. Treatment of RAW264.7 cells with CpG oligodeoxynucleotide (ODN) for both 18 and 42 h resulted in a significant increase in DNA binding, whereas non-CpG ODN had no effect on DNA binding. Enhanced DNA binding was non-sequence-specific, inhibited by unlabeled DNA, showed saturation, was consistent with increased cell surface DNA receptors, and resulted in enhanced internalization of DNA. Treatment with Escherichia coli DNA or lipopolysaccharide (LPS) also resulted in a significant increase in DNA binding, but treatment with interleukin-1α, tumor necrosis factor-α, or phorbol 12-myristate 13-acetate had no effect on DNA binding. Soluble factors produced in response to treatment with CpG ODN or LPS did not affect DNA binding. These studies demonstrate that one consequence of activating the host innate immune response by bacterial infection is enhanced binding and internalization of DNA. During this period of increased DNA internalization, RAW264.7 cells were hypo-responsive to continued stimulation by CpG ODN, as assessed by tumor necrosis factor-α activity. We speculate the biological significance of increasing DNA binding and internalization following interaction with bacterial PAMPs may provide a mechanism to limit an ongoing immune inflammatory response by enhancing clearance of bacterial DNA from the extracellular environment.",
author = "McCoy, {Sharon L.} and Kurtz, {Stephen E.} and Hausman, {Frances A.} and Dennis Trune and Bennett, {Robert (Rob)} and Steven Hefeneider",
year = "2004",
month = "4",
day = "23",
doi = "10.1074/jbc.M303837200",
language = "English (US)",
volume = "279",
pages = "17217--17223",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "17",

}

TY - JOUR

T1 - Activation of RAW264.7 Macrophages by Bacterial DNA and Lipopolysaccharide Increases Cell Surface DNA Binding and Internalization

AU - McCoy, Sharon L.

AU - Kurtz, Stephen E.

AU - Hausman, Frances A.

AU - Trune, Dennis

AU - Bennett, Robert (Rob)

AU - Hefeneider, Steven

PY - 2004/4/23

Y1 - 2004/4/23

N2 - Bacterial DNA containing unmethylated CpG motifs is a pathogen-associated molecular pattern (PAMP) that interacts with host immune cells via a toll-like receptor (TLR) to induce immune responses. DNA binding and internalization into cells is independent of TLR expression, receptor-mediated, and required for cell activation. The objective of this study was to determine whether exposure of immune cells to bacterial DNA affects DNA binding and internalization. Treatment of RAW264.7 cells with CpG oligodeoxynucleotide (ODN) for both 18 and 42 h resulted in a significant increase in DNA binding, whereas non-CpG ODN had no effect on DNA binding. Enhanced DNA binding was non-sequence-specific, inhibited by unlabeled DNA, showed saturation, was consistent with increased cell surface DNA receptors, and resulted in enhanced internalization of DNA. Treatment with Escherichia coli DNA or lipopolysaccharide (LPS) also resulted in a significant increase in DNA binding, but treatment with interleukin-1α, tumor necrosis factor-α, or phorbol 12-myristate 13-acetate had no effect on DNA binding. Soluble factors produced in response to treatment with CpG ODN or LPS did not affect DNA binding. These studies demonstrate that one consequence of activating the host innate immune response by bacterial infection is enhanced binding and internalization of DNA. During this period of increased DNA internalization, RAW264.7 cells were hypo-responsive to continued stimulation by CpG ODN, as assessed by tumor necrosis factor-α activity. We speculate the biological significance of increasing DNA binding and internalization following interaction with bacterial PAMPs may provide a mechanism to limit an ongoing immune inflammatory response by enhancing clearance of bacterial DNA from the extracellular environment.

AB - Bacterial DNA containing unmethylated CpG motifs is a pathogen-associated molecular pattern (PAMP) that interacts with host immune cells via a toll-like receptor (TLR) to induce immune responses. DNA binding and internalization into cells is independent of TLR expression, receptor-mediated, and required for cell activation. The objective of this study was to determine whether exposure of immune cells to bacterial DNA affects DNA binding and internalization. Treatment of RAW264.7 cells with CpG oligodeoxynucleotide (ODN) for both 18 and 42 h resulted in a significant increase in DNA binding, whereas non-CpG ODN had no effect on DNA binding. Enhanced DNA binding was non-sequence-specific, inhibited by unlabeled DNA, showed saturation, was consistent with increased cell surface DNA receptors, and resulted in enhanced internalization of DNA. Treatment with Escherichia coli DNA or lipopolysaccharide (LPS) also resulted in a significant increase in DNA binding, but treatment with interleukin-1α, tumor necrosis factor-α, or phorbol 12-myristate 13-acetate had no effect on DNA binding. Soluble factors produced in response to treatment with CpG ODN or LPS did not affect DNA binding. These studies demonstrate that one consequence of activating the host innate immune response by bacterial infection is enhanced binding and internalization of DNA. During this period of increased DNA internalization, RAW264.7 cells were hypo-responsive to continued stimulation by CpG ODN, as assessed by tumor necrosis factor-α activity. We speculate the biological significance of increasing DNA binding and internalization following interaction with bacterial PAMPs may provide a mechanism to limit an ongoing immune inflammatory response by enhancing clearance of bacterial DNA from the extracellular environment.

UR - http://www.scopus.com/inward/record.url?scp=2342423630&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=2342423630&partnerID=8YFLogxK

U2 - 10.1074/jbc.M303837200

DO - 10.1074/jbc.M303837200

M3 - Article

C2 - 14757773

AN - SCOPUS:2342423630

VL - 279

SP - 17217

EP - 17223

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 17

ER -