Atomic Model of Rabbit Hemorrhagic Disease Virus by Cryo-Electron Microscopy and Crystallography

Xue Wang,Klaus Schulten,Fengting Xu,Jun Ma,Yujia Zhai,Hai Pang,Yanxin Liu,Kai Zhang,Jiasen Liu,Fei Sun,Timothy S Baker,Dong Zheng,Bingquan Gao,Erica Ollmann Saphire

doi:10.1371/journal.ppat.1003132

Abstract

Rabbit hemorrhagic disease, first described in China in 1984, causes hemorrhagic necrosis of the liver. Its etiological agent, rabbit hemorrhagic disease virus (RHDV), belongs to the Lagovirus genus in the family Caliciviridae. The detailed molecular structure of any lagovirus capsid has yet to be determined. Here, we report a cryo-electron microscopic (cryoEM) reconstruction of wild-type RHDV at 6.5 Å resolution and the crystal structures of the shell (S) and protruding (P) domains of its major capsid protein, VP60, each at 2.0 Å resolution. From these data we built a complete atomic model of the RHDV capsid. VP60 has a conserved S domain and a specific P2 sub-domain that differs from those found in other caliciviruses. As seen in the shell portion of the RHDV cryoEM map, which was resolved to ∼5.5 Å, the N-terminal arm domain of VP60 folds back onto its cognate S domain. Sequence alignments of VP60 from six groups of RHDV isolates revealed seven regions of high variation that could be mapped onto the surface of the P2 sub-domain and suggested three putative pockets might be responsible for binding to histo-blood group antigens. A flexible loop in one of these regions was shown to interact with rabbit tissue cells and contains an important epitope for anti-RHDV antibody production. Our study provides a reliable, pseudo-atomic model of a Lagovirus and suggests a new candidate for an efficient vaccine that can be used to protect rabbits from RHDV infection.

Highlights

Rabbit hemorrhagic disease (RHD) is extremely contagious in adult rabbits and is often associated with liver necrosis, hemorrhaging, and high mortality [1]
We find that rabbit hemorrhagic disease virus (RHDV) VP60 has a P2 sub-domain that differs from other caliciviruses
Concluding remarks In this study, we used cryo-electron microscopic (cryoEM) methods to reconstruct the structure of the RHDV capsid to an overall estimated resolution limit of 6.5 A (5.5 Ain the shell domain) and solved the crystal structures of the S and P domains of the RHDV VP60 protein both at 2.0 Aresolution

Summary

Introduction

Rabbit hemorrhagic disease (RHD) is extremely contagious in adult rabbits and is often associated with liver necrosis, hemorrhaging, and high mortality [1]. RHD outbreaks still occur on almost every continent and cause significant mortality rates, being endemic in Europe, Asia, Africa, and Australia [4] This disease has a significant impact on the rabbit industry and ecology [4]. Previous structural studies of caliciviruses include threedimensional (3D) cryo-electron microscopic (cryoEM) reconstructions of virus-like particles (VLPs) of Murine Norovirus (MNV, Norovirus) and Feline calicivirus (FCV, Vesivirus) at 8- and 16-Aresolution, respectively [10,11], and determination of the crystal structures of the Norwalk virus (NV, Norovirus) capsid at 3.4 A [12], native FCV virions at 3.6 A [13], and native virions of San Miguel sea lion virus (SMSV, Vesivirus) at 3.2 A [14]. CryoEM reconstructions of the RHDV VLP at 8 A [15] and the native RHDV virion at 11 A [7] have been computed and a Ca homology model of RHDV was built based on

Author Summary

Findings

Materials and Methods