Cellular Localization and Antigenic Characterization of Crimean-Congo Hemorrhagic Fever Virus Glycoproteins

Abstract

Crimean-Congo hemorrhagic fever virus (CCHFV), a member of the genus Nairovirus of the family Bunyaviridae , causes severe disease with high rates of mortality in humans. The CCHFV M RNA segment encodes the virus glycoproteins G _N and G _C . To understand the processing and intracellular localization of the CCHFV glycoproteins as well as their neutralization and protection determinants, we produced and characterized monoclonal antibodies (MAbs) specific for both G _N and G _C . Using these MAbs, we found that G _N predominantly colocalized with a Golgi marker when expressed alone or with G _C , while G _C was transported to the Golgi apparatus only in the presence of G _N . Both proteins remained endo-β- N -acetylglucosaminidase H sensitive, indicating that the CCHFV glycoproteins are most likely targeted to the cis Golgi apparatus. Golgi targeting information partly resides within the G _N ectodomain, because a soluble version of G _N lacking its transmembrane and cytoplasmic domains also localized to the Golgi apparatus. Coexpression of soluble versions of G _N and G _C also resulted in localization of soluble G _C to the Golgi apparatus, indicating that the ectodomains of these proteins are sufficient for the interactions needed for Golgi targeting. Finally, the mucin-like and P35 domains, located at the N terminus of the G _N precursor protein and removed posttranslationally by endoproteolysis, were required for Golgi targeting of G _N when it was expressed alone but were dispensable when G _C was coexpressed. In neutralization assays on SW-13 cells, MAbs to G _C , but not to G _N , prevented CCHFV infection. However, only a subset of G _C MAbs protected mice in passive-immunization experiments, while some nonneutralizing G _N MAbs efficiently protected animals from a lethal CCHFV challenge. Thus, neutralization of CCHFV likely depends not only on the properties of the antibody, but on host cell factors as well. In addition, nonneutralizing antibody-dependent mechanisms, such as antibody-dependent cell-mediated cytotoxicity, may be involved in the in vivo protection seen with the MAbs to G _C .