Cryo-electron microscopy reconstructions of two types of wild rabbit hemorrhagic disease viruses characterized the structural features of Lagovirus

Rabbit hemorrhagic disease was described in China in 1984 and can cause hemorrhagic necrosis of the liver within two or three days after infection.The etiological agent,rabbit hemorrhagic disease virus(RHDV),belongs to the Lagovirus genus in the Caliciviridae family.Compared to other calicivirus,such as rNV and SMSV,the structure of Lagovirus members is not well characterized.In this report,structures of two types of wild RHDV particles,the intact virion and the core-like particle(CLP),were reconstructed by cryo-electron microscopy at 11Åand 17Å,respectively.This is the first time the 3D structure of wild caliciviruses CLP has been provided,and the 3D structure of intact RHDV virion is the highest resolution structure in Lagovirus.Comparison of the intact virion and CLP structures clearly indicated that CLP was produced from the intact virion with the protrusion dissociated.In contrast with the crystal structures of recombinant Norovirus and San Miguel sea lion virus,the capsomers of RHDV virion exhibited unique structural features and assembly modes.Both P1 and P2 subdomains have interactions inside the AB capsomer,while only P2 subdomains have interaction inside CC capsomer.The pseudo atomic models of RHDV capsomers were constructed by homology modeling and density map fitting,and the rotation of RHDV VP60 P domain with respect to its S domain,compared with SMSV,was observed.Collectively,our cryo-electron microscopic studies of RHDV provide close insight into the structure of Lagovirus,which is important for functional analysis and better vaccine development in the future.

Nucleolin interacts with the rabbit hemorrhagic disease virus replicase RdRp, nonstructural proteins p16 and p23, playing a role in virus replication

Rabbit hemorrhagic disease virus(RHDV)is a member of the Caliciviridae family and cannot be propagated in vitro,which has impeded the progress of investigating its replication mechanism.Construction of an RHDV replicon system has recently provided a platform for exploring RHDV replication in host cells.Here,aided by this replicon system and using two-step affinity purification,we purified the RHDV replicase and identified its associated host factors.We identified rabbit nucleolin(NCL)as a physical link,which mediating the interaction between other RNA-dependent RNA polymerase(Rd Rp)-related host proteins and the viral replicase Rd Rp.We found that the overexpression or knockdown of NCL significantly increased or severely impaired RHDV replication in RK-13 cells,respectively.NCL was identified to directly interact with RHDV Rd Rp,p16,and p23.Furthermore,NCL knockdown severely impaired the binding of Rd Rp to Rd Rp-related host factors.Collectively,these results indicate that the host protein NCL is essential for RHDV replication and acts as a physical link between viral replicase and host proteins.