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Streptococcus pneumoniae stabilizes tumor necrosis factor alpha mRNA through a pathway dependent on p38 MAPK but independent of Toll-like receptors.

Mogensen TH, Berg RS, Ostergaard L, Paludan SR - BMC Immunol. (2008)

Bottom Line: The ability of S. pneumoniae to stabilize TNF-alpha mRNA was dependent on the mitogen-activated protein kinase (MAPK) p38 whereas inhibition of Toll-like receptor signaling via MyD88 did not affect S. pneumoniae-induced mRNA stabilization.P38 was activated through a pathway involving the upstream kinase transforming growth factor-activated kinase 1 and MAPK kinase 3.Production of TNF-alpha may contribute significantly to the inflammatory response raised during pneumococcal infection.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Infectious Diseases, Skejby Hospital - Aarhus University Hospital, DK-8200, Aarhus N, Denmark. trine.mogensen@dadlnet.dk

ABSTRACT

Background: Streptococcus pneumoniae is a human pathogenic bacteria and a major cause of severe invasive diseases, including pneumonia, bacteremia, and meningitis. Infections with S. pneumoniae evoke a strong inflammatory response, which plays a major role in the pathogenesis of pneumococcal disease.

Results: In this study, we have examined how S. pneumoniae affects expression of the inflammatory cytokine tumor necrosis factor (TNF) alpha, and the molecular mechanisms involved. Secretion of TNF-alpha was strongly induced by S. pneumoniae, which was able to stabilize TNF-alpha mRNA through a mechanism dependent on the viability of the bacteria as well as the adenylate uridylate-rich elements in the 3'untranslated region of TNF-alpha mRNA. The ability of S. pneumoniae to stabilize TNF-alpha mRNA was dependent on the mitogen-activated protein kinase (MAPK) p38 whereas inhibition of Toll-like receptor signaling via MyD88 did not affect S. pneumoniae-induced mRNA stabilization. P38 was activated through a pathway involving the upstream kinase transforming growth factor-activated kinase 1 and MAPK kinase 3.

Conclusion: Thus, S. pneumoniae stabilizes TNF-alpha mRNA through a pathway dependent on p38 but independent of Toll-like receptors. Production of TNF-alpha may contribute significantly to the inflammatory response raised during pneumococcal infection.

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P38 MAPK is important for stabilization of TNF-α mRNA by S. pneumoniae. (A) RAW-TNF-AU+ cells were treated with the inhibitors SB202190 (p38 and RIP2) (5 μM) and PP2 (Src and RIP2) (500 nM) 15 min prior to addition of vehicle or S. pneumoniae (5 × 107 bacteria/ml) to the cell cultures. Total cell lysates were harvested 20 h later and CAT was measured by ELISA. The data is shown as means +/- SEM. (B) RAW264.7 cells were treated with live or heat-killed S. pneumoniae (5 × 107 bacteria/ml) for the indicated amount of time, and whole cell lysates were isolated. Phosphorylation of p38 was measured by Luminex. The data is shown as means +/- SEM. (C and D) RAW264.7 (C) and C57BL/6 macrophages (D) were incubated with a MyD88 inhibitor peptide or a control peptide for 24 h before addition of vehicle or S. pneumoniae (5 × 107 bacteria/ml). Whole cell lysates were harvested 30 min later and phosphorylation of p38 was measured by Luminex. The data is shown as means +/- SEM. Similar results were obtained in 2–3 independent experiments.
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Figure 4: P38 MAPK is important for stabilization of TNF-α mRNA by S. pneumoniae. (A) RAW-TNF-AU+ cells were treated with the inhibitors SB202190 (p38 and RIP2) (5 μM) and PP2 (Src and RIP2) (500 nM) 15 min prior to addition of vehicle or S. pneumoniae (5 × 107 bacteria/ml) to the cell cultures. Total cell lysates were harvested 20 h later and CAT was measured by ELISA. The data is shown as means +/- SEM. (B) RAW264.7 cells were treated with live or heat-killed S. pneumoniae (5 × 107 bacteria/ml) for the indicated amount of time, and whole cell lysates were isolated. Phosphorylation of p38 was measured by Luminex. The data is shown as means +/- SEM. (C and D) RAW264.7 (C) and C57BL/6 macrophages (D) were incubated with a MyD88 inhibitor peptide or a control peptide for 24 h before addition of vehicle or S. pneumoniae (5 × 107 bacteria/ml). Whole cell lysates were harvested 30 min later and phosphorylation of p38 was measured by Luminex. The data is shown as means +/- SEM. Similar results were obtained in 2–3 independent experiments.

Mentions: The signaling pathways responsible for mediating mRNA stability have recently been studied extensively, and it appears that P38 MAPK plays a central role in this process [13]. To examine the role of p38 in stabilization of TNF-α mRNA in response to S. pneumoniae infection, we treated the RAW TNF-α 3'-UTR AU+ cell line with the p38 inhibitor SB202190 15 min prior to addition of S. pneumoniae. CAT was subsequently measured in cell lysates. As shown in Fig. 4A, the strong elevation of CAT protein levels observed after infection with live S. pneumoniae was significantly inhibited by the presence of SB202190. It has been reported that the p38 MAPK inhibitor SB203580, which is structurally very similar to SB202190, not only inhibits p38 but also the kinase receptor-interacting protein (RIP) 2 [27], which plays an important role in signaling downstream of the S. pneumoniae-activated PRR NOD2 [27]. Therefore, we also examined the effect of another RIP2 inhibitor, PP2 [27], on induction of CAT protein in our reporter system However, as illustrated in Fig. 4A, no effect of PP2 treatment towards bacteria-induced CAT production was observed.


Streptococcus pneumoniae stabilizes tumor necrosis factor alpha mRNA through a pathway dependent on p38 MAPK but independent of Toll-like receptors.

Mogensen TH, Berg RS, Ostergaard L, Paludan SR - BMC Immunol. (2008)

P38 MAPK is important for stabilization of TNF-α mRNA by S. pneumoniae. (A) RAW-TNF-AU+ cells were treated with the inhibitors SB202190 (p38 and RIP2) (5 μM) and PP2 (Src and RIP2) (500 nM) 15 min prior to addition of vehicle or S. pneumoniae (5 × 107 bacteria/ml) to the cell cultures. Total cell lysates were harvested 20 h later and CAT was measured by ELISA. The data is shown as means +/- SEM. (B) RAW264.7 cells were treated with live or heat-killed S. pneumoniae (5 × 107 bacteria/ml) for the indicated amount of time, and whole cell lysates were isolated. Phosphorylation of p38 was measured by Luminex. The data is shown as means +/- SEM. (C and D) RAW264.7 (C) and C57BL/6 macrophages (D) were incubated with a MyD88 inhibitor peptide or a control peptide for 24 h before addition of vehicle or S. pneumoniae (5 × 107 bacteria/ml). Whole cell lysates were harvested 30 min later and phosphorylation of p38 was measured by Luminex. The data is shown as means +/- SEM. Similar results were obtained in 2–3 independent experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
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Figure 4: P38 MAPK is important for stabilization of TNF-α mRNA by S. pneumoniae. (A) RAW-TNF-AU+ cells were treated with the inhibitors SB202190 (p38 and RIP2) (5 μM) and PP2 (Src and RIP2) (500 nM) 15 min prior to addition of vehicle or S. pneumoniae (5 × 107 bacteria/ml) to the cell cultures. Total cell lysates were harvested 20 h later and CAT was measured by ELISA. The data is shown as means +/- SEM. (B) RAW264.7 cells were treated with live or heat-killed S. pneumoniae (5 × 107 bacteria/ml) for the indicated amount of time, and whole cell lysates were isolated. Phosphorylation of p38 was measured by Luminex. The data is shown as means +/- SEM. (C and D) RAW264.7 (C) and C57BL/6 macrophages (D) were incubated with a MyD88 inhibitor peptide or a control peptide for 24 h before addition of vehicle or S. pneumoniae (5 × 107 bacteria/ml). Whole cell lysates were harvested 30 min later and phosphorylation of p38 was measured by Luminex. The data is shown as means +/- SEM. Similar results were obtained in 2–3 independent experiments.
Mentions: The signaling pathways responsible for mediating mRNA stability have recently been studied extensively, and it appears that P38 MAPK plays a central role in this process [13]. To examine the role of p38 in stabilization of TNF-α mRNA in response to S. pneumoniae infection, we treated the RAW TNF-α 3'-UTR AU+ cell line with the p38 inhibitor SB202190 15 min prior to addition of S. pneumoniae. CAT was subsequently measured in cell lysates. As shown in Fig. 4A, the strong elevation of CAT protein levels observed after infection with live S. pneumoniae was significantly inhibited by the presence of SB202190. It has been reported that the p38 MAPK inhibitor SB203580, which is structurally very similar to SB202190, not only inhibits p38 but also the kinase receptor-interacting protein (RIP) 2 [27], which plays an important role in signaling downstream of the S. pneumoniae-activated PRR NOD2 [27]. Therefore, we also examined the effect of another RIP2 inhibitor, PP2 [27], on induction of CAT protein in our reporter system However, as illustrated in Fig. 4A, no effect of PP2 treatment towards bacteria-induced CAT production was observed.

Bottom Line: The ability of S. pneumoniae to stabilize TNF-alpha mRNA was dependent on the mitogen-activated protein kinase (MAPK) p38 whereas inhibition of Toll-like receptor signaling via MyD88 did not affect S. pneumoniae-induced mRNA stabilization.P38 was activated through a pathway involving the upstream kinase transforming growth factor-activated kinase 1 and MAPK kinase 3.Production of TNF-alpha may contribute significantly to the inflammatory response raised during pneumococcal infection.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Infectious Diseases, Skejby Hospital - Aarhus University Hospital, DK-8200, Aarhus N, Denmark. trine.mogensen@dadlnet.dk

ABSTRACT

Background: Streptococcus pneumoniae is a human pathogenic bacteria and a major cause of severe invasive diseases, including pneumonia, bacteremia, and meningitis. Infections with S. pneumoniae evoke a strong inflammatory response, which plays a major role in the pathogenesis of pneumococcal disease.

Results: In this study, we have examined how S. pneumoniae affects expression of the inflammatory cytokine tumor necrosis factor (TNF) alpha, and the molecular mechanisms involved. Secretion of TNF-alpha was strongly induced by S. pneumoniae, which was able to stabilize TNF-alpha mRNA through a mechanism dependent on the viability of the bacteria as well as the adenylate uridylate-rich elements in the 3'untranslated region of TNF-alpha mRNA. The ability of S. pneumoniae to stabilize TNF-alpha mRNA was dependent on the mitogen-activated protein kinase (MAPK) p38 whereas inhibition of Toll-like receptor signaling via MyD88 did not affect S. pneumoniae-induced mRNA stabilization. P38 was activated through a pathway involving the upstream kinase transforming growth factor-activated kinase 1 and MAPK kinase 3.

Conclusion: Thus, S. pneumoniae stabilizes TNF-alpha mRNA through a pathway dependent on p38 but independent of Toll-like receptors. Production of TNF-alpha may contribute significantly to the inflammatory response raised during pneumococcal infection.

Show MeSH
Related in: MedlinePlus