The C, V and W proteins of Nipah virus inhibit minigenome replication

Abstract

Nipah virus (NiV) is a recently emergent, highly pathogenic, zoonotic paramyxovirus of the genus Henipavirus. Like the phosphoprotein (P) gene of other paramyxoviruses, the P gene of NiV is predicted to encode three additional proteins, C, V and W. When the C, V and W proteins of NiV were tested for their ability to inhibit expression of the chloramphenicol acetyltransferase (CAT) reporter gene in plasmid-based, minigenome replication assays, each protein inhibited CAT expression in a dose-dependent manner. The C, V and W proteins of NiV also inhibited expression of CAT from a measles virus (MV) minigenome, but not from a human parainfluenzavirus 3 (hPIV3) minigenome. Interestingly, the C and V proteins of MV, which have previously been shown to inhibit MV minigenome replication, also inhibited NiV minigenome replication; however, they were not able to inhibit hPIV3 minigenome replication. In contrast, the C protein of hPIV3 inhibited minigenome replication of hPIV3, NiV and MV. Although there is very limited amino acid sequence similarity between the C, V and W proteins within the paramyxoviruses, the heterotypic inhibition of replication suggests that these proteins may share functional properties. INTRODUCTIONNipah virus (NiV) is a recently emergent, highly pathogenic, zoonotic paramyxovirus. NiV was first identified in Malaysia in 1998 during an outbreak of neurological and respiratory disease in swine, which resulted in severe febrile encephalitis in humans who had contact with infected pigs (Chua et al., 2000). NiV has been associated with subsequent outbreaks of fatal, febrile encephalitis in Bangladesh and India between (Hsu et al., 2004Luby et al., 2006). Molecular characterization of NiV revealed that it was related closely to Hendra virus, another zoonotic paramyxovirus, which had emerged in Australia in 1994 (Chua et al., 2000;Harcourt et al., 2000Harcourt et al., , 2001. These two viruses constitute the recently recognized genus Henipavirus (Mayo, 2002).The genome of NiV is 18 246 nt in length and contains six transcription units encoding six viral structural proteins (39-N-P-M-F-G-L-59) and three predicted non-structural proteins, C, V and W . As in other paramyxoviruses, the C protein of NiV is expressed from an alternative open reading frame (ORF) within the phosphoprotein (P) gene, whereas the V and W proteins are expressed by RNA editing (Lamb & Kolakofsky, 2001). At a unique, highly conserved RNA-editing site in the P gene, the polymerase protein (L) inserts a single, nontemplated G residue that results in a frame shift and the expression of the V protein. Insertion of two nontemplated G residues results in expression of the W protein . Whilst the C proteins of NiV and of other paramyxoviruses are unique and share no sequence similarity with the P protein (Lamb & Kolakofsky, 2001), the V and W proteins share an amino-terminal 407 aa domain with P and each possesses a unique carboxyl-terminal domain consisting of 52 aa for V and 47 aa for W . The sequences of the P proteins of paramyxoviruses a...

Determination of the henipavirus phosphoprotein gene mRNA editing frequencies and detection of the C, V and W proteins of Nipah virus in virus-infected cells

“…The W protein was detected exclusively in the nucleus and was distributed evenly throughout, with the exception of the nucleoli. The cellular distributions of V and W proteins were consistent with previous studies in which the V and W ORFs were individually expressed via plasmids (Rodriguez & Horvath, 2004;Rodriguez et al, 2002;Shaw et al, 2004Shaw et al, , 2005.The C, V and W proteins of NiV inhibit both transcription and replication in a NiV minigenome assay (Sleeman et al, 2008). The distinct cellular distributions of each protein shown in this study imply that the inhibitory effects of these proteins utilize different mechanisms.…”

“…The C, V and W proteins of NiV inhibit both transcription and replication in a NiV minigenome assay (Sleeman et al, 2008). The distinct cellular distributions of each protein shown in this study imply that the inhibitory effects of these proteins utilize different mechanisms.…”

Efficient Reverse Genetics Reveals Genetic Determinants of Budding and Fusogenic Differences between Nipah and Hendra Viruses and Enables Real-Time Monitoring of Viral Spread in Small Animal Models of Henipavirus Infection

“…The HHrbz sequence in stem I designed to anneal to the NiV 3= leader, TTGGT, was mutated to TCGGT so as to match the HeV 3= leader. For the support plasmids, HeV-N, -P, and -L were cloned into pTM1 as described previously for NiV (33), with the two C ORF start codons mutated in P (from ATGATG to ACGACG) to prevent potential inhibition of rescue (34).…”

Nonstructural Nipah Virus C Protein Regulates both the Early Host Proinflammatory Response and Viral Virulence

“…The NiV C protein has also been shown to inhibit the activation of an antiviral state, but the mechanism is unknown (43). Together those studies, as well as experiments showing that the C, V, and W proteins can inhibit replication of a minigenome (52), have been performed using plasmid-based expression analysis. Finally, using the recombinant NiV, lacking the expression of accessory proteins, NiV V and C were shown to be important in viral virulence, but in contrast to the other paramyxoviruses, none of the three NiV nonstructural proteins seemed to affect interferon signaling in infected cells (63), leaving the mechanism of their action unclear.…”