Characterization of Schistosoma japonicum CP1412 protein as a novel member of the ribonuclease T2 molecule family with immune regulatory function.
Characterization of Schistosoma japonicum CP1412 protein as a novel member of the ribonuclease T2 molecule family with immune regulatory function.
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DOI：
10.1186/s13071-016-1962-y
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发表时间：
2017-02-17
影响因子：
3.2
通讯作者：
Zhou W
中科院分区：
文献类型：
--
作者：
Ke XD;Shen S;Song LJ;Yu CX;Kikuchi M;Hirayama K;Gao H;Wang J;Yin X;Yao Y;Liu Q;Zhou W
血吸虫感染通常诱导极化的Th 2型宿主免疫应答。由于虫卵抗原分子在血吸虫病免疫调节过程中起关键作用，阐明其在血吸虫病中的功能将有助于疫苗和免疫方法的发展。日本血吸虫CP 1412（GenBank：AY57074.1）是核糖核酸酶T2家族的新成员，具有免疫调节功能。构建了重组表达质粒Sj CP 1412-pET 28 a，转化大肠杆菌，经IPTG诱导表达重组蛋白（rSj CP 1412）。利用生物信息学方法预测了Sj CP 1412的RNase活性，并通过rSj CP 1412消化酵母tRNA的实验进行了验证。同时，观察了Sj CP 1412的RNase活性与免疫调节的关系。 Sj CP 1412是一种新的具有RNase活性的RNase T2家族蛋白。免疫印迹和RT-PCR分析表明，Sj CP 1412作为一种蛋白质，专门分泌/排泄的卵，但不是尾蚴和蠕虫。rSj CP 1412刺激RAW264.7巨噬细胞后，CD 206、Arg-1和IL-10表达增加，与M2型巨噬细胞分化有关。用rSjCP 1412刺激树突状细胞（DC）未能诱导其成熟，并且重组蛋白也抑制LPS刺激的DC成熟。从可溶性卵抗原（SEA）中耗尽Sj CP 1412削弱SEA诱导RAW 264.7巨噬细胞M2型极化的能力。rSj CP 1412免疫小鼠，可诱导小鼠产生高滴度的抗体，提高血清IL-4、TGF-β水平和脾脏CD 4 + CD 25 + Foxp 3 + T细胞水平，下调血清IFN-γ水平，并减轻虫卵肉芽肿病理。rSj CP 1412的体外刺激显著增加健康小鼠脾细胞中的CD 4 + CD 25 + Foxp 3 + T细胞数量。经DEPC灭活的具有RNase活性的rSj CP 1412蛋白不能诱导RAW 264.7巨噬细胞表达M2表面标志物CD 206。Sj CP 1412蛋白在S.日本血吸虫卵是RNase T2家族的新成员。与曼氏血吸虫的Omega-1类似，Sj CP 1412蛋白驱动宿主Th 2免疫应答的极化，这取决于其RNA酶活性。这些数据为理解核糖核酸酶T2家族蛋白在病毒感染过程中的免疫调节作用提供了新的证据。本文的在线版本（doi：10.1186/s13071-016-1962-y）包含补充材料，可供授权用户使用。
Schistosome infection typically induces a polarized Th2 type host immune response. As egg antigen molecules play key roles in this immunoregulatory process, clarifying their functions in schistosomiasis would facilitate the development of vaccine and immunotherapeutic methods. Schistosoma japonicum (Sj) CP1412 (GenBank: AY57074.1) has been identified as a new member of the RNase T2 family with immune regulatory functions. The expression plasmid Sj CP1412-pET28a was constructed and transformed into bacteria for production of recombinant Sj CP1412 protein (rSj CP1412) via IPTG induction. The RNase activity of Sj CP1412 was predicted by bioinformatic analysis and confirmed by digesting the yeast tRNA with rSj CP1412.C57BL/6j mice were immunized with rSj CP1412, and its immune regulatory effects in vivo and in vitro were investigated. Meanwhile, the relationship between the RNase activity of Sj CP1412 and its immune regulation was observed. Sj CP1412 was confirmed as a novel RNase T2 family protein with RNase activity. Immunoblotting and RT-PCR analyses demonstrated Sj CP1412 as a protein exclusively secreted/excreted from eggs, but not cercariae and adult worms. Stimulating RAW264.7 macrophages with rSj CP1412 raised the expression of CD206, Arg-1 and IL-10, which are related to M2 type macrophage differentiation. Stimulating dendritic cells (DCs) with rSjCP1412 failed to induce their maturation, and the recombinant protein also inhibited LPS-stimulated DC maturation. Depletion of Sj CP1412 from soluble egg antigen (SEA) impaired the ability of SEA to induce M2 type polarization of RAW264.7 macrophages. Immunizing mice with rSj CP1412 induced high antibody titers, increased serum IL-4 and TGF-β levels and splenic CD4 + CD25 + Foxp3 + T cells, downregulated serum IFN-γ levels and alleviated the egg granuloma pathology of schistosome infection. In vitro stimulation by rSj CP1412 significantly increased CD4 + CD25 + Foxp3 + T cell numbers in splenocytes of healthy mice. The rSj CP1412 protein with RNase activity inactivated by DEPC failed to induce M2 surface marker CD206 expression in RAW264.7 macrophages. The Sj CP1412 protein expressed specifically in S. japonicum eggs is a novel member of the RNase T2 family. Similar to Omega-1 of Schistosoma mansoni, the Sj CP1412 protein drives polarization of the host Th2 immune response, which is dependent on its RNase activity. These data provide new evidence towards understanding the immune regulatory role of RNase T2 family proteins during schistosome infection. The online version of this article (doi:10.1186/s13071-016-1962-y) contains supplementary material, which is available to authorized users.