Characterization of Schistosoma japonicum CP1412 protein as a novel member of the ribonuclease T2 molecule family with immune regulatory function

Abstract

BackgroundSchistosome 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.MethodsThe 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.Results 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.ConclusionsThe 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.Electronic supplementary materialThe online version of this article (doi:10.1186/s13071-016-1962-y) contains supplementary material, which is available to authorized users.

Modulation of Host Immunity by Helminths: The Expanding Repertoire of Parasite Effector Molecules

“…Omega-1 does not degrade specific transcripts, but rather targets the global pool of RNA. In this way omega-1, and its close homolog from Schistosoma japonicum , CP1412, suppress DC activation and maturation, preventing upregulation of CD86 and MHCII and synthesis of IL-12 in response to CD40 ligation, favoring both Th2 cell ( Everts et al., 2012 , Everts et al., 2009 , Steinfelder et al., 2009 , Wilbers et al., 2017 ) and regulatory T (Treg) cell ( Ke et al., 2017 , Zaccone et al., 2011 ) induction. T cell priming in the absence of IL-12 and with diminished co-stimulation may favor Th2 cell polarization.…”

Alterations of Gut Microbiome and Metabolite Profiling in Mice Infected by Schistosoma japonicum

“…Different stages of S. japonicum cause various damages to the host, and complex immune pathological reactions lead to diverse clinical symptoms. Larval worms induce Th1 responses with elevated levels of the inflammatory cytokines IFN-γ, IL-12, and TNF-α in the early phase of schistosomiasis and cause diarrhea, fatigue, and anemia, while adult worms become mature and lay eggs; parasite eggs that deposit in the liver and colon of infected hosts elicit Th2 responses and then upregulate the serum cytokine levels of IL-4, IL-5, IL-13, and TGF-β, leading to portal vein hypertension syndrome, ascites, and liver fibrosis ( 2 , 4 , 5 ). During S. japonicum infection, egg deposits in the tissues are a determining factor that shifts the Th1 response to the Th2 response ( 5 ).…”

Programmed genome editing of the omega-1 ribonuclease 1 of the blood fluke,Schistosoma mansoni

“…The ribonuclease activity of the ω1 glycoprotein in SEA is associated with the Th2-polarized immune response that underpins the appearance of schistosome egg granulomata (19, 22). Ribonuclease activity of SEA from control and experimental groups on substrate yeast RNA was investigated following CRISPR/Cas9 programmed mutation of ω1 mediated by the RNP and the pseudotyped lentiviral approaches with or without ssODN (20, 50). Intact yeast RNA was evident in the DNase-RNase free condition (negative control), indicative of absence of RNase activity in the reagents (200 ng yeast RNA at the outset).…”

“…The RNase activity of ω1 in wild type SEA or Δω1-SEA was analyzed by visualizing and quantifying the substrate that remained following enzymolysis by agarose gel electrophoresis and staining with ethidium bromide. The yeast RNA digestion by control SEAs or Δω1-SEA were set up in triplicates, with quantity of residual RNA determined by densitometry [48].…”