Staphylococcal peptidoglycan co-localizes with Nod2 and TLR2 and activates innate immune response via both receptors in primary murine keratinocytes

Maria Anna Müller-Anstett, Patrick Müller, Till Albrecht, Mulugeta Nega, Jeanette Wagener, Qiang Gao, Susanne Kaesler, Martin Schaller, Tilo Biedermann, Friedrich Götz

Research output: Contribution to journalArticle

56 Citations (Scopus)
3 Downloads (Pure)

Abstract

In mammalian host cells staphylococcal peptidoglycan (PGN) is recognized by Nod2. Whether PGN is also recognized by TLR2 is disputed. Here we carried out PGN co-localization and stimulation studies with TLR2 and Nod2 in wild type and mutant host cells. To exclude contamination with lipoproteins, polymeric staphylococcal PGN (PGN(pol)) was isolated from Staphylococcus aureus ¿lgt (lacking lipidated prelipoproteins). PGN(pol) was biotinylated (PGN-Bio) for fluorescence monitoring with specific antibodies. Keratinocytes from murine oral epithelium (MK) readily internalized PGN-Bio in an endocytosis-like process. In wt MK, PGN(pol) induced intracellular accumulation of Nod2 and TLR2 and co-localized with Nod2 and TLR2, but not with TLR4. In TLR2-deficient MK Nod2 and in Nod2-deficient MK TLR2 was induced, indicating that PGN(pol) recognition by Nod2 is independent of TLR2 and vice versa. In both mutants IL-6 and IL-1B release was decreased by approximately 50% compared to wt MK, suggesting that the immune responses induced by Nod2 and TLR2 are comparable and that the two receptors act additively in MK. In TLR2-transfected HEK293 cells PGN(pol) induced NFkB-promoter fused luciferase expression. To support the data, co-localization and signaling studies were carried out with SHL-PGN, a lipase protein covalently tethered to PGN-fragments of varying sizes at its C-terminus. SHL-PGN also co-localized with Nod2 or TLR2 and induced their accumulation, while SHL without PGN did not. The results show that staphylococcal PGN not only co-localizes with Nod2 but also with TLR2. PGN is able to stimulate the immune system via both receptors.
Original languageEnglish
Article numbere13153
Number of pages13
JournalPloS ONE
Volume5
Issue number10
DOIs
Publication statusPublished - 7 Oct 2010

Fingerprint

Peptidoglycan
peptidoglycans
keratinocytes
Keratinocytes
Innate Immunity
receptors
mice
innate immunity
mutants
Environmental Monitoring
HEK293 Cells
Immune system
endocytosis
cells
luciferase
Endocytosis
Luciferases
Lipase

Keywords

  • amino acid sequence
  • animals
  • biotin
  • electrophoresis, polyacrylamide gel
  • endocytosis
  • immunity, innate
  • keratinocytes
  • mice
  • molecular sequence data
  • mouth mucosa
  • NF-kappa B
  • Nod2 signaling adaptor protein
  • peptidoglycan
  • spectrometry, fluorescence
  • Staphylococcus aureus
  • toll-like receptor 2

Cite this

Staphylococcal peptidoglycan co-localizes with Nod2 and TLR2 and activates innate immune response via both receptors in primary murine keratinocytes. / Müller-Anstett, Maria Anna; Müller, Patrick; Albrecht, Till; Nega, Mulugeta; Wagener, Jeanette; Gao, Qiang; Kaesler, Susanne; Schaller, Martin; Biedermann, Tilo; Götz, Friedrich.

In: PloS ONE, Vol. 5, No. 10, e13153, 07.10.2010.

Research output: Contribution to journalArticle

Müller-Anstett, MA, Müller, P, Albrecht, T, Nega, M, Wagener, J, Gao, Q, Kaesler, S, Schaller, M, Biedermann, T & Götz, F 2010, 'Staphylococcal peptidoglycan co-localizes with Nod2 and TLR2 and activates innate immune response via both receptors in primary murine keratinocytes', PloS ONE, vol. 5, no. 10, e13153. https://doi.org/10.1371/journal.pone.0013153
Müller-Anstett, Maria Anna ; Müller, Patrick ; Albrecht, Till ; Nega, Mulugeta ; Wagener, Jeanette ; Gao, Qiang ; Kaesler, Susanne ; Schaller, Martin ; Biedermann, Tilo ; Götz, Friedrich. / Staphylococcal peptidoglycan co-localizes with Nod2 and TLR2 and activates innate immune response via both receptors in primary murine keratinocytes. In: PloS ONE. 2010 ; Vol. 5, No. 10.
@article{d6c6584948b246bda9f190d512e779f8,
title = "Staphylococcal peptidoglycan co-localizes with Nod2 and TLR2 and activates innate immune response via both receptors in primary murine keratinocytes",
abstract = "In mammalian host cells staphylococcal peptidoglycan (PGN) is recognized by Nod2. Whether PGN is also recognized by TLR2 is disputed. Here we carried out PGN co-localization and stimulation studies with TLR2 and Nod2 in wild type and mutant host cells. To exclude contamination with lipoproteins, polymeric staphylococcal PGN (PGN(pol)) was isolated from Staphylococcus aureus ¿lgt (lacking lipidated prelipoproteins). PGN(pol) was biotinylated (PGN-Bio) for fluorescence monitoring with specific antibodies. Keratinocytes from murine oral epithelium (MK) readily internalized PGN-Bio in an endocytosis-like process. In wt MK, PGN(pol) induced intracellular accumulation of Nod2 and TLR2 and co-localized with Nod2 and TLR2, but not with TLR4. In TLR2-deficient MK Nod2 and in Nod2-deficient MK TLR2 was induced, indicating that PGN(pol) recognition by Nod2 is independent of TLR2 and vice versa. In both mutants IL-6 and IL-1B release was decreased by approximately 50{\%} compared to wt MK, suggesting that the immune responses induced by Nod2 and TLR2 are comparable and that the two receptors act additively in MK. In TLR2-transfected HEK293 cells PGN(pol) induced NFkB-promoter fused luciferase expression. To support the data, co-localization and signaling studies were carried out with SHL-PGN, a lipase protein covalently tethered to PGN-fragments of varying sizes at its C-terminus. SHL-PGN also co-localized with Nod2 or TLR2 and induced their accumulation, while SHL without PGN did not. The results show that staphylococcal PGN not only co-localizes with Nod2 but also with TLR2. PGN is able to stimulate the immune system via both receptors.",
keywords = "amino acid sequence, animals, biotin, electrophoresis, polyacrylamide gel, endocytosis, immunity, innate, keratinocytes, mice, molecular sequence data, mouth mucosa, NF-kappa B, Nod2 signaling adaptor protein, peptidoglycan, spectrometry, fluorescence, Staphylococcus aureus, toll-like receptor 2",
author = "M{\"u}ller-Anstett, {Maria Anna} and Patrick M{\"u}ller and Till Albrecht and Mulugeta Nega and Jeanette Wagener and Qiang Gao and Susanne Kaesler and Martin Schaller and Tilo Biedermann and Friedrich G{\"o}tz",
year = "2010",
month = "10",
day = "7",
doi = "10.1371/journal.pone.0013153",
language = "English",
volume = "5",
journal = "PloS ONE",
issn = "1932-6203",
publisher = "PUBLIC LIBRARY SCIENCE",
number = "10",

}

TY - JOUR

T1 - Staphylococcal peptidoglycan co-localizes with Nod2 and TLR2 and activates innate immune response via both receptors in primary murine keratinocytes

AU - Müller-Anstett, Maria Anna

AU - Müller, Patrick

AU - Albrecht, Till

AU - Nega, Mulugeta

AU - Wagener, Jeanette

AU - Gao, Qiang

AU - Kaesler, Susanne

AU - Schaller, Martin

AU - Biedermann, Tilo

AU - Götz, Friedrich

PY - 2010/10/7

Y1 - 2010/10/7

N2 - In mammalian host cells staphylococcal peptidoglycan (PGN) is recognized by Nod2. Whether PGN is also recognized by TLR2 is disputed. Here we carried out PGN co-localization and stimulation studies with TLR2 and Nod2 in wild type and mutant host cells. To exclude contamination with lipoproteins, polymeric staphylococcal PGN (PGN(pol)) was isolated from Staphylococcus aureus ¿lgt (lacking lipidated prelipoproteins). PGN(pol) was biotinylated (PGN-Bio) for fluorescence monitoring with specific antibodies. Keratinocytes from murine oral epithelium (MK) readily internalized PGN-Bio in an endocytosis-like process. In wt MK, PGN(pol) induced intracellular accumulation of Nod2 and TLR2 and co-localized with Nod2 and TLR2, but not with TLR4. In TLR2-deficient MK Nod2 and in Nod2-deficient MK TLR2 was induced, indicating that PGN(pol) recognition by Nod2 is independent of TLR2 and vice versa. In both mutants IL-6 and IL-1B release was decreased by approximately 50% compared to wt MK, suggesting that the immune responses induced by Nod2 and TLR2 are comparable and that the two receptors act additively in MK. In TLR2-transfected HEK293 cells PGN(pol) induced NFkB-promoter fused luciferase expression. To support the data, co-localization and signaling studies were carried out with SHL-PGN, a lipase protein covalently tethered to PGN-fragments of varying sizes at its C-terminus. SHL-PGN also co-localized with Nod2 or TLR2 and induced their accumulation, while SHL without PGN did not. The results show that staphylococcal PGN not only co-localizes with Nod2 but also with TLR2. PGN is able to stimulate the immune system via both receptors.

AB - In mammalian host cells staphylococcal peptidoglycan (PGN) is recognized by Nod2. Whether PGN is also recognized by TLR2 is disputed. Here we carried out PGN co-localization and stimulation studies with TLR2 and Nod2 in wild type and mutant host cells. To exclude contamination with lipoproteins, polymeric staphylococcal PGN (PGN(pol)) was isolated from Staphylococcus aureus ¿lgt (lacking lipidated prelipoproteins). PGN(pol) was biotinylated (PGN-Bio) for fluorescence monitoring with specific antibodies. Keratinocytes from murine oral epithelium (MK) readily internalized PGN-Bio in an endocytosis-like process. In wt MK, PGN(pol) induced intracellular accumulation of Nod2 and TLR2 and co-localized with Nod2 and TLR2, but not with TLR4. In TLR2-deficient MK Nod2 and in Nod2-deficient MK TLR2 was induced, indicating that PGN(pol) recognition by Nod2 is independent of TLR2 and vice versa. In both mutants IL-6 and IL-1B release was decreased by approximately 50% compared to wt MK, suggesting that the immune responses induced by Nod2 and TLR2 are comparable and that the two receptors act additively in MK. In TLR2-transfected HEK293 cells PGN(pol) induced NFkB-promoter fused luciferase expression. To support the data, co-localization and signaling studies were carried out with SHL-PGN, a lipase protein covalently tethered to PGN-fragments of varying sizes at its C-terminus. SHL-PGN also co-localized with Nod2 or TLR2 and induced their accumulation, while SHL without PGN did not. The results show that staphylococcal PGN not only co-localizes with Nod2 but also with TLR2. PGN is able to stimulate the immune system via both receptors.

KW - amino acid sequence

KW - animals

KW - biotin

KW - electrophoresis, polyacrylamide gel

KW - endocytosis

KW - immunity, innate

KW - keratinocytes

KW - mice

KW - molecular sequence data

KW - mouth mucosa

KW - NF-kappa B

KW - Nod2 signaling adaptor protein

KW - peptidoglycan

KW - spectrometry, fluorescence

KW - Staphylococcus aureus

KW - toll-like receptor 2

U2 - 10.1371/journal.pone.0013153

DO - 10.1371/journal.pone.0013153

M3 - Article

VL - 5

JO - PloS ONE

JF - PloS ONE

SN - 1932-6203

IS - 10

M1 - e13153

ER -