Dataset Information

Microneme Protein 6 Is Involved in Invasion and Egress by Neospora caninum.

ABSTRACT:

Background

Neospora caninum, is the etiological agent of neosporosis, an infection that causes abortions in cattle and nervous system dysfunction in dogs. Invasion and egress are the key steps of the pathogenesis of N. caninum infection. Microneme proteins (MICs) play important roles in the recognition, adhesion, and invasion of host cells in other apicomplexan parasites. However, some MICs and their functions in N. caninum infection have rarely been reported.

Methods

The homologous recombination strategy was used to investigate the function of MIC6 in N. caninum infection.

Results

?NcMIC6 showed a smaller plaque size and weakened capacities of invasion and egress than Nc1. Transcription levels of the egress-related genes CDPK1, PLP1, and AMA1 of ?NcMIC6 were downregulated. Due to the lack of NcMIC6, virulence of the pathogen in the infected mouse was weakened. The subcellular localization of NcMIC1 and NcMIC4 in ?NcMIC6, however, did not change. Nevertheless, the transcription levels of MIC1 and MIC4 in ?NcMIC6 were downregulated, and the expression and secretion of MIC1 and MIC4 in ?NcMIC6 were reduced compared with that in Nc1. Furthermore, the absence of NcMIC6 weakened the virulence in mice and lower parasite load detected in mice brains.

Conclusions

NcMIC6 is involved in host cell invasion and egress in N. caninum and may work synergistically with other MICs to regulate the virulence of the pathogen. These data lay a foundation for further research into the function and application of NcMIC6.

SUBMITTER: Wang X

PROVIDER: S-EPMC7918358 | biostudies-literature | 2021 Feb

REPOSITORIES: biostudies-literature

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Publications

Microneme Protein 6 Is Involved in Invasion and Egress by <i>Neospora caninum</i>.

Wang Xianmei X Tang Di D Wang Fei F Jin Gaowei G Wang Lifang L Liu Qun Q Liu Jing J

Pathogens (Basel, Switzerland) 20210213 2

<h4>Background</h4><i>Neospora caninum</i>, is the etiological agent of neosporosis, an infection that causes abortions in cattle and nervous system dysfunction in dogs. Invasion and egress are the key steps of the pathogenesis of <i>N. caninum</i> infection. Microneme proteins (MICs) play important roles in the recognition, adhesion, and invasion of host cells in other apicomplexan parasites. However, some MICs and their functions in <i>N. caninum</i> infection have rarely been reported.<h4>Met ...[more]

PMID: 33668497

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Project description:The invasive stages of apicomplexan parasites enter their host cells through mechanisms which are largely conserved throughout the phylum. Host cell invasion is divided into two distinct events, namely, adhesion onto the host cell surface and the actual host cell entry process. The former is mediated largely through microneme proteins which are secreted at the onset of establishing contact with the host cell surface. Many of the microneme proteins identified so far contain adhesive domains. We here present the genomic and corresponding cDNA sequences coding for a 460-amino-acid (aa) microneme protein in Neospora caninum tachyzoites which, due to its homology to MIC1 in Toxoplasma gondii (TgMIC1), was named NcMIC1. The deduced NcMIC1 polypeptide sequence contains an N-terminal signal peptide of 20 aa followed by two tandemly internal repeats of 48 and 44 aa, respectively. Integrated into each repeat is a CXXXCG sequence motif reminiscent of the thrombospondin-related family of adhesive proteins. The positioning of this motif is strictly conserved in TgMIC1 and NcMIC1. The C-terminal part, comprised of 278 aa, was expressed in Escherichia coli, and antibodies affinity purified on recombinant NcMIC1 were used to confirm the localization within the micronemes by immunofluorescence and immunogold transmission electron microscopy of tachyzoites. Immunohistochemistry of mouse brains infected with tissue cysts showed that expression of this protein is reduced in the bradyzoite stage. Upon initiation of secretion by elevating the temperature to 37 degrees C, NcMIC1 is released into the medium supernatant. NcMIC1 binds to trypsinized, rounded Vero cells, as well as to Vero cell monolayers. Removal of glycosaminoglycans from the host cell surface and modulation of host cell surface glycosaminoglycan sulfation significantly reduces the binding of NcMIC1 to the host cell surface. Solid-phase binding assays employing defined glycosaminoglycans confirmed that NcMIC1 binds to sulfated glycosaminoglycans.