Dataset Information

14-3-3 Protein of Neospora caninum Modulates Host Cell Innate Immunity Through the Activation of MAPK and NF-?B Pathways.

ABSTRACT: Neospora caninum is an obligate intracellular apicomplexan parasite, the etiologic agent of neosporosis, and a major cause of reproductive loss in cattle. There is still a lack of effective prevention and treatment measures. The 14-3-3 protein is a widely expressed acidic protein that spontaneously forms dimers within apicomplexan parasites. This protein has been isolated and sequenced in many parasites; however, there are few reports about the N. caninum 14-3-3 protein. Here, we successfully expressed and purified a recombinant fusion protein of Nc14-3-3 (rNc14-3-3) and prepared a polyclonal antibody. Immunofluorescence and immunogold electron microscopy studies of tachyzoites or N. caninum-infected cells suggested that 14-3-3 was localized in the cytosol and the membrane. Western blotting analysis indicated that rNc14-3-3 could be recognized by N. caninum-infected mouse sera, suggesting that 14-3-3 may be an infection-associated antigen that is involved in the host immune response. We demonstrated that rNc14-3-3 induced cytokine expression by activating the MAPK and AKT signaling pathways, and inhibitors of p38, ERK, JNK, and AKT could significantly decrease the production of IL-6, IL-12p40, and TNF-?. In addition, phosphorylated nuclear factor-?B (NF-?B/p65) was observed in wild-type peritoneal macrophages (PMs) treated with rNc14-3-3, and the protein level of NF-?B/p65 was reduced in the cytoplasm but increased correspondingly in the nucleus after 2 h of treatment. These results were also observed in deficient in TLR2-/- PMs. Taken together, our results indicated that the N. caninum 14-3-3 protein can induce effective immune responses and stimulate cytokine expression by activating the MAPK, AKT, and NF-?B signaling pathways but did not dependent TLR2, suggesting that Nc14-3-3 is a novel vaccine candidate against neosporosis.

SUBMITTER: Li S

PROVIDER: S-EPMC6355710 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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14-3-3 Protein of Neospora caninum Modulates Host Cell Innate Immunity Through the Activation of MAPK and NF-κB Pathways.

Li Shan S Gong Pengtao P Zhang Nan N Li Xin X Tai Lixin L Wang Xu X Yang Zhengtao Z Yang Ju J Zhu Xingquan X Zhang Xichen X Li Jianhua J

Frontiers in microbiology 20190125

Neospora caninum is an obligate intracellular apicomplexan parasite, the etiologic agent of neosporosis, and a major cause of reproductive loss in cattle. There is still a lack of effective prevention and treatment measures. The 14-3-3 protein is a widely expressed acidic protein that spontaneously forms dimers within apicomplexan parasites. This protein has been isolated and sequenced in many parasites; however, there are few reports about the N. caninum 14-3-3 protein. Here, we s ...[more]

PMID: 30740096

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Project description:Neospora caninum is an obligate intracellular parasite, which causes significant economic losses in the cattle industry. However, the immune mechanism of the parasite-host interaction is not yet fully understood. Extracellular vesicles (EVs) have emerged as a ubiquitous mechanism by which almost all cells, especially immune and tumor cells, participate in intercellular communications. Although studies have indicated that EVs secreted by Toxoplasma gondii or Trypanosoma brucei promote exchanges of biological molecules important for the host-parasite interplay, however, EVs and their biological activities in N. caninum is not clear. Here, we used multiple methods, including electron microscopy, nanoparticle tracking analysis, RT-PCR, immunofluorescence, western blot, proteomics, and cytokine analyses, to examine the properties of N. caninum EVs. We found that N. caninum produced EVs that are similar to mammalian exosomes, which generally range from 30 to 150 nm in diameter. It was shown that N. caninum EVs could remarkably increase the production of pro-inflammatory cytokines IL-12p40, TNF-α, IL-1β, IL-6, and IFN-γ by wild-type (WT) mouse bone marrow-derived macrophages (BMDMs) whereas the secretion of IL-12p40, TNF-α, and IFN-γ was very strongly downregulated in TLR2-/- mouse BMDMs. The levels of IL-6 were not affected, but the secretion of IL-10 was upregulated. We found that the phosphorylation levels of P38, ERK, and JNK were significantly reduced in the TLR2-/- cells compared with those in WT mouse BMDMs and that treatment with chemical inhibiters of P38, ERK, and JNK resulted in upregulation of IL-6, IL-12p40, and IL-10 production. Together, these results demonstrated that N. caninum EVs could be rapidly internalized to deliver proteins to the host cells and modulate the host cell immune responses through MAPK signaling pathway in a TLR2-dependent manner. Our study is the first to reveal potential roles for N. caninum EVs in host communication and immune response in parasite-host interactions.

Dataset Information

14-3-3 Protein of Neospora caninum Modulates Host Cell Innate Immunity Through the Activation of MAPK and NF-?B Pathways.

Publications

14-3-3 Protein of <i>Neospora caninum</i> Modulates Host Cell Innate Immunity Through the Activation of MAPK and NF-κB Pathways.

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