CryoEM analysis of the Nipah virus ribonucleoprotein complex

The Nipah virus (NiV) is a highly pathogenic RNA virus and a member of the Paramyxoviridae family. Its genomic RNA is wrapped by the nucleocapsid (N) protein in a helical manner forming a long flexible ribonucleoprotein (RNP) assembly. This assembly serves to protect the viral genome from ribonuclease degradation and the host immune response. The RNP assembly also serves as a functional template for the viral RNA synthesis by the synergistic action of the viral RNA dependent RNA polymerase (RdRp) complex which made up of viral large RNA polymerase (L protein) and the viral cofactor phosphoprotein (P protein). However, the molecular details of this viral RNA synthesis remain elusive. This research aims to better understand the initial step of viral RNA synthesis by elucidating the molecular interaction of the NiV RNP assembly and its viral cofactor P protein. The bacterial expression of the NiV nucleocapsid protein led to the formation of the nucleocapsid-like helical assembly containing bacterial cellular RNA. The NiV nucleocapsid protein-RNA complex was purified and its structure was determined using cryo electron microscopy (cryoEM). The structure reveals the nonspecific binding of the RNA in a “three-bases-in, three-bases-out” conformation by the nucleocapsid protein, as well as the role of the N- and C-terminal segments of the nucleocapsid protein in stabilising the RNP assembly. A stable protein complex between the NiV RNP assembly and the P protein was prepared using bacterial co-expression systems. The cryoEM analysis of the NiV N/P heterocomplex suggests the uncoiling of the RNP helical assembly upon interaction with the P protein. Structural data on the NiV nucleocapsid assembly reported here provide molecular insight into the assembly and structure of the NiV RNP complexes and will guide further work towards understanding the mechanisms of viral RNA synthesis.