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TLR4 mediates pneumolysin-induced ATF3 expression through the JNK/p38 pathway in Streptococcus pneumoniae-infected RAW 264.7 cells.

Nguyen CT, Kim EH, Luong TT, Pyo S, Rhee DK - Mol. Cells (2014)

Bottom Line: The expression of ATF3 was induced by pneumolysin (PLY), a potent pneumococcal virulence factor, via the TLR4 pathway.Furthermore, ATF3 induction is mediated by p38 mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase (JNK).Thus, this study reveals a potential role of PLY in modulating ATF3 expression, which is required for the regulation of immune responses against pneumococcal infection in macrophages.

View Article: PubMed Central - PubMed

Affiliation: School of Pharmacy, Sungkyunkwan University, Suwon 440-746, Korea.

ABSTRACT
Activating transcription factor-3 (ATF3) acts as a negative regulator of cytokine production during Gram-negative bacterial infection. A recent study reported that ATF3 provides protection from Streptococcus pneumoniae infection by activating cytokines. However, the mechanism by which S. pneumoniae induces ATF3 after infection is still unknown. In this study, we show that ATF3 was upregulated via Toll-like receptor (TLR) pathways in response to S. pneumoniae infection in vitro. Induction was mediated by TLR4 and TLR2, which are in the TLR family. The expression of ATF3 was induced by pneumolysin (PLY), a potent pneumococcal virulence factor, via the TLR4 pathway. Furthermore, ATF3 induction is mediated by p38 mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase (JNK). Thus, this study reveals a potential role of PLY in modulating ATF3 expression, which is required for the regulation of immune responses against pneumococcal infection in macrophages.

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The TLR2/4 pathway is required for activation of JNK/p38. RAW 264.7 cells were transfected with siTLR4 (A) or siTLR2 (B) for 24 h and then infected with S. pneumoniae for 0, 1, 2, or 4 h. Subsequently, the cell lysates were analyzed by Western blotting. The band densities were measured by ImageJ. The data are representative of three independent experiments and were analyzed by one-way ANOVA. Data show mean ± s.d. *P < 0.05, **P < 0.01, ***P < 0.001.
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f2-molcell-38-1-58: The TLR2/4 pathway is required for activation of JNK/p38. RAW 264.7 cells were transfected with siTLR4 (A) or siTLR2 (B) for 24 h and then infected with S. pneumoniae for 0, 1, 2, or 4 h. Subsequently, the cell lysates were analyzed by Western blotting. The band densities were measured by ImageJ. The data are representative of three independent experiments and were analyzed by one-way ANOVA. Data show mean ± s.d. *P < 0.05, **P < 0.01, ***P < 0.001.

Mentions: In pneumococcal infection, MAPKs play an important role in generating innate immune responses and production of mediators in macrophages (Kang et al., 2009). The results obtained clearly show that phosphorylation of p38 and ERK was increased by pneumococcal infection 10 min post-infection (Figs. 2A and 2B), whereas the phosphorylation of JNK was significantly induced 30 min post-infection (Figs. 2A and 2B). Interestingly, siTLR4 treatment slightly reduced the induction of p-JNK at 30 min and 60 min post-infection, whereas treatment with siTLR2 significantly abolished the levels of p-JNK expression (Figs. 2A and 2B). Moreover, the phosphorylation of p38 markedly decreased in both siTLR4- and siTLR2-transfected cells. However, the phosphorylation of ERK was not TLR4- or TLR2-dependent (Figs. 2A and 2B). Taken together, these results indicate that S. pneumoniae-induced phosphorylation of JNK and p38 is TLR2/4-dependent.


TLR4 mediates pneumolysin-induced ATF3 expression through the JNK/p38 pathway in Streptococcus pneumoniae-infected RAW 264.7 cells.

Nguyen CT, Kim EH, Luong TT, Pyo S, Rhee DK - Mol. Cells (2014)

The TLR2/4 pathway is required for activation of JNK/p38. RAW 264.7 cells were transfected with siTLR4 (A) or siTLR2 (B) for 24 h and then infected with S. pneumoniae for 0, 1, 2, or 4 h. Subsequently, the cell lysates were analyzed by Western blotting. The band densities were measured by ImageJ. The data are representative of three independent experiments and were analyzed by one-way ANOVA. Data show mean ± s.d. *P < 0.05, **P < 0.01, ***P < 0.001.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4314132&req=5

f2-molcell-38-1-58: The TLR2/4 pathway is required for activation of JNK/p38. RAW 264.7 cells were transfected with siTLR4 (A) or siTLR2 (B) for 24 h and then infected with S. pneumoniae for 0, 1, 2, or 4 h. Subsequently, the cell lysates were analyzed by Western blotting. The band densities were measured by ImageJ. The data are representative of three independent experiments and were analyzed by one-way ANOVA. Data show mean ± s.d. *P < 0.05, **P < 0.01, ***P < 0.001.
Mentions: In pneumococcal infection, MAPKs play an important role in generating innate immune responses and production of mediators in macrophages (Kang et al., 2009). The results obtained clearly show that phosphorylation of p38 and ERK was increased by pneumococcal infection 10 min post-infection (Figs. 2A and 2B), whereas the phosphorylation of JNK was significantly induced 30 min post-infection (Figs. 2A and 2B). Interestingly, siTLR4 treatment slightly reduced the induction of p-JNK at 30 min and 60 min post-infection, whereas treatment with siTLR2 significantly abolished the levels of p-JNK expression (Figs. 2A and 2B). Moreover, the phosphorylation of p38 markedly decreased in both siTLR4- and siTLR2-transfected cells. However, the phosphorylation of ERK was not TLR4- or TLR2-dependent (Figs. 2A and 2B). Taken together, these results indicate that S. pneumoniae-induced phosphorylation of JNK and p38 is TLR2/4-dependent.

Bottom Line: The expression of ATF3 was induced by pneumolysin (PLY), a potent pneumococcal virulence factor, via the TLR4 pathway.Furthermore, ATF3 induction is mediated by p38 mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase (JNK).Thus, this study reveals a potential role of PLY in modulating ATF3 expression, which is required for the regulation of immune responses against pneumococcal infection in macrophages.

View Article: PubMed Central - PubMed

Affiliation: School of Pharmacy, Sungkyunkwan University, Suwon 440-746, Korea.

ABSTRACT
Activating transcription factor-3 (ATF3) acts as a negative regulator of cytokine production during Gram-negative bacterial infection. A recent study reported that ATF3 provides protection from Streptococcus pneumoniae infection by activating cytokines. However, the mechanism by which S. pneumoniae induces ATF3 after infection is still unknown. In this study, we show that ATF3 was upregulated via Toll-like receptor (TLR) pathways in response to S. pneumoniae infection in vitro. Induction was mediated by TLR4 and TLR2, which are in the TLR family. The expression of ATF3 was induced by pneumolysin (PLY), a potent pneumococcal virulence factor, via the TLR4 pathway. Furthermore, ATF3 induction is mediated by p38 mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase (JNK). Thus, this study reveals a potential role of PLY in modulating ATF3 expression, which is required for the regulation of immune responses against pneumococcal infection in macrophages.

Show MeSH
Related in: MedlinePlus