Submitted to: Experimental Parasitology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/1/2010
Publication Date: 5/21/2010
Citation: Jenkins, M.C., Tuo, W., Feng, X., Cao, L., Murphy, C.A., Fetterer, R.H. 2010. Neospora caninum: Cloning and expression of a gene coding for cytokine-inducing Neospora caninum profilin. Experimental Parasitology. 125:357-362. Interpretive Summary: Neospora caninum is a protozoan that is a major cause of reproductive failure in dairy cattle worldwide. Although a number of subunit vaccines have been tested, none elicits complete protection against experimental N. caninum challenge. One possible reason for this lack of protective immunity is that important immune responses are not being induced that are relevant in a natural infection. In the present study, an antigen termed NcProfilin was produced in harmless Escherichia coli using recombinant DNA technology. Using antibodies to the recombinant protein, NcProfilin was localized to the surface and secreted by tachyzoite stages of the parasite. In addition, this protein stimulated the release of immune cytokines thought to be involved in protective immunity against N. caninum infection. Thus, NcProfilin represents a potential vaccine candidate that may be used alone or in conjunction with other recombinant N. caninum proteins to stimulate protection against neosporosis.
Technical Abstract: Profilins are actin-binding proteins that in T. gondii stimulate innate immunity in mice by binding Toll-like receptors (TLR) on dendritic cells (DC) leading to release of inflammatory cytokines, primarily IL-12 and IFN-'. The purpose of the present study was to characterize Neospora caninum profilin, termed NcProfilin, by expressing the cDNA sequence in Escherichia coli. Recombinant NcProfilin was purified by affinity chromatography, and used to prepare specific antisera to characterize native NcProfilin antigen in N. caninum tachyzoites. By immunoblotting, recombinant NcProfilin is 21 kDa, and is similar in size to the respective 22 kDa native protein. Immunofluorescence and immuno-electron microscopy revealed native NcProfilin to localize to the apical end of N. caninum tachyzoites. Methanol treatment of tachyzoites prior to immuno-fluorescence staining, revealed a surface or subsurface locale. Incubation of recombinant NcProfilin with spleen cells or enriched dendritic cells from BALB/c mice induced release of IL-12, IFN-', and TNF-a.