Structural and functional characterization of Reston Ebola VP35 Interferon Inhibitory Domain

摘要

Ebola viruses (EBOV) are causative agents of lethal hemorrhagic fever in humans and non-human primates. Among the filoviruses characterized thus far, Reston Ebola virus (REBOV) is the only Ebola species that is non-pathogenic in humans despite the fact that REBOV can cause lethal disease in non-human primates. Previous studies also suggest that Reston EBOV is less effective at inhibiting host innate immune responses, compared with Zaire EBOV or Marburg virus. Virally encoded VP35 protein is critical for immune suppression, but an understanding of the relative contributions of VP35 proteins from REBOV and other filoviruses is currently lacking. In order to address this question, we characterized REBOV VP35 IFN inhibitory domain (IID) using structural, biochemical, and virological studies. These studies reveal differences in dsRNA binding and IFN inhibition between the two species. These observed differences are likely due to increased stability and loss of flexibility in REBOV VP35 IID as demonstrated by thermal-shift stability assays. Consistent with this finding, our 1.71 Å crystal structure of the REBOV VP35 IID reveal that the structure is highly similar to ZEBOV VP35 IID with an overall backbone r.m.s.d. of 0.64 Å, but contains an additional helical element at the linker between the two subdomains of VP35 IID. Mutations near the linker, including swapping sequences between REBOV and ZEBOV, reveal that the linker sequence has limited tolerance for variability. Together with the previously solved ligand free and dsRNA bound forms of ZEBOV VP35 IID structures, our current studies of REBOV VP35 IID reinforce the importance of VP35 in immune suppression. Functional differences observed between REBOV and ZEBOV VP35 proteins may contribute to observed differences in pathogenicity, but these are unlikely be the major determinant. However, the high similarity in structure and the low tolerance of sequence variability, coupled with the multiple critical roles played by EBOV VP35 proteins, highlight the viability of VP35 as a potential target for therapeutic development.