Eimeria tenella Eimeria-specific protein that interacts with apical membrane antigen 1 (EtAMA1) is involved in host cell invasion
Eimeria tenella Eimeria-specific protein that interacts with apical membrane antigen 1 (EtAMA1) is involved in host cell invasion
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柔嫩艾美耳球虫 与顶膜抗原 1 (EtAMA1) 相互作用的艾美耳球虫特异性蛋白参与宿主细胞侵袭
DOI：
10.1186/s13071-020-04229-5
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发表时间：
2020-07-25
影响因子：
3.2
通讯作者：
Han, Hongyu
中科院分区：
文献类型：
--
作者：
Li, Cong;Zhao, Qiping;Han, Hongyu
研究背景鸡球虫病是一种广泛分布的、经济上具有重要意义的禽类疾病，由几种艾美耳球虫引起。这些寄生虫具有复杂多样的生命周期，需要入侵其宿主细胞。这是由顶端分泌细胞器分泌的各种蛋白质介导的。顶膜抗原1（AMA1），这是从微丝释放，并在所有顶复门保守，在宿主细胞入侵中起着核心作用。在以前的研究中，一些假定的ETAMA1相互作用蛋白的E。tenella进行筛选。在这项研究中，我们确定了一个假定的EtAMA1相互作用蛋白，E。方法采用双分子荧光互补技术（BiFC）和谷胱甘肽S转移酶（GST）融合蛋白下拉技术（GST pull-down）研究EtAMA1与EtEspin在体内和体外的相互作用，并分析EtEspin在E. tenella进行定量PCR和Western blotting。当子孢子在41 ° C完全培养基中孵育时，用星形孢菌素测试EtEsp蛋白的分泌。用免疫荧光法（IFA）分析EtEsp的定位。体外侵袭抑制实验检测抗EtEsp抗体对E.结果EtAMA1和EtEsp之间的相互作用通过BiFC和GST pull-down证实。我们的结果表明EtEsp在寄生虫生命周期的不同阶段有差异表达。免疫荧光分析表明，EtEsp蛋白主要分布在寄生虫的表面，并且随着寄生虫在宿主细胞中的发育，该蛋白的表达量逐渐增加。利用星形孢菌素，我们发现EtEsp是一种分泌蛋白，但不是来自微丝。在抑制试验中，多克隆抗rEtEsp抗体减弱了E.结论EtEsp与EtAMA1相互作用，该蛋白为分泌蛋白，而不是来自微线体。该蛋白参与子孢子对宿主细胞的侵入，并可能参与寄生虫的生长。这些数据对使用EtAMA1或EtAMA1相互作用蛋白作为针对禽球虫病的干预策略的靶点具有意义。
BackgroundAvian coccidiosis is a widespread, economically significant disease of poultry, caused by severalEimeriaspecies. These parasites have complex and diverse life-cycles that require invasion of their host cells. This is mediated by various proteins secreted from apical secretory organelles. Apical membrane antigen 1 (AMA1), which is released from micronemes and is conserved across all apicomplexans, plays a central role in the host cell invasion. In a previous study, some putativeEtAMA1-interacting proteins ofE. tenellawere screened. In this study, we characterized one putativeEtAMA1-interacting protein,E. tenella Eimeria-specific protein (EtEsp).MethodsBimolecular fluorescence complementation (BiFC) and glutathione S-transferase (GST) fusion protein pull-down (GST pull-down) were used to confirm the interaction betweenEtAMA1 andEtEspin vivoandin vitro.The expression ofEtEsp was analyzed in different developmental stages ofE. tenellawith quantitative PCR and western blotting. The secretion ofEtEsp protein was tested with staurosporine when sporozoites were incubated in complete medium at 41 °C. The localization ofEtEsp was analyzed with an immunofluorescence assay (IFA). Anin vitroinvasion inhibition assay was conducted to assess the ability of antibodies againstEtEsp to inhibit cell invasion byE. tenellasporozoites.ResultsThe interaction betweenEtAMA1 andEtEsp was confirmed with BiFC and by GST pull-down. Our results show thatEtEsp is differentially expressed during distinct phases of the parasite life-cycle. IFA showed that theEtEsp protein is mainly distributed on the parasite surface, and that the expression of this protein increases during the development of the parasite in the host cells. Using staurosporine, we showed thatEtEsp is a secreted protein, but not from micronemes. In inhibition tests, a polyclonal anti-rEtEsp antibody attenuated the capacity ofE. tenellato invade host cells.ConclusionIn this study, we show thatEtEsp interacts withEtAMA1 and that the protein is secreted protein, but not from micronemes. The protein participates in sporozoite invasion of host cells and is maybe involved in the growth of the parasite. These data have implications for the use ofEtAMA1 orEtAMA1-interacting proteins as targets in intervention strategies against avian coccidiosis.