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Listeriolysin O-Dependent Bacterial Entry into the Cytoplasm Is Required for Calpain Activation and Interleukin-1α Secretion in Macrophages Infected with Listeria monocytogenes
- Dewamitta, Sita R., Nomura, Takamasa, Kawamura, Ikuo, Hara, Hideki, Tsuchiya, Kohsuke, Kurenuma, Takeshi, Shen, Yanna, Daim, Sylvia, Yamamoto, Takeshi, Qu, Huixin, Sakai, Shunsuke, Xu, Yanting, Mitsuyama, Masao
- Infection and immunity 2010 v.78 no.5 pp. 1884-1894
- Listeria monocytogenes, bacteria, calcium, calcium signaling, calpain, chelating agents, cytokines, cytoplasm, genes, immune response, macrophages, messenger RNA, mutants, secretion
- Listeriolysin O (LLO), an hly-encoded cytolysin of Listeria monocytogenes, plays an essential role in the entry of L. monocytogenes into the host cell cytoplasm. L. monocytogenes-infected macrophages produce various proinflammatory cytokines, including interleukin-1α (IL-1α), that contribute to the host immune response. In this study, we have examined IL-1α production in macrophages infected with wild-type L. monocytogenes or a nonescaping mutant strain deficient for LLO (Δhly). Expression of IL-1α mRNA and accumulation of pro-IL-1α in the cytoplasm were induced by both strains. In contrast, the secretion of the mature form of IL-1α from infected macrophages was observed in infection with wild-type L. monocytogenes but not with the Δhly mutant. A recovery of the ability to induce IL-1α secretion was shown in a mutant strain complemented with the hly gene. The Toll-like receptor (TLR)/MyD88 signaling pathway was exclusively required for the expression of pro-IL-1α, independently of LLO-mediated cytoplasmic entry of L. monocytogenes. The LLO-dependent secretion of mature IL-1α was abolished by addition of calcium chelators, and only LLO-producing L. monocytogenes strains were able to induce elevation of the intracellular calcium level in infected macrophages. A calcium-dependent protease, calpain, was implicated in the maturation and secretion of IL-1α induced by LLO-producing L. monocytogenes strains based on the effect of calpain inhibitor. Functional activation of calpain was detected in macrophages infected with LLO-producing L. monocytogenes strains but not with a mutant strain lacking LLO. These results clearly indicated that LLO-mediated cytoplasmic entry of bacteria could induce the activation of intracellular calcium signaling, which is essential for maturation and secretion of IL-1α in macrophages during L. monocytogenes infection through activation of a calcium-dependent calpain protease. In addition, recombinant LLO, when added to macrophages infected with the Δhly strain, could induce calcium influx and IL-1α secretion at doses exhibiting cytolytic activity, suggesting that LLO produced by intracellular L. monocytogenes may be implicated in induction of calcium influx through pore formation.