Nidovirus RNA polymerases: Complex enzymes handling exceptional RNA genomes

Abstract

Coronaviruses and arteriviruses are distantly related human and animal pathogens that belong to the order Nidovirales. Nidoviruses are characterized by their polycistronic plus-stranded RNA genome, the production of subgenomic mRNAs and the conservation of a specific array of replicase domains, including key RNA-synthesizing enzymes. Coronaviruses (26-34 kilobases) have the largest known RNA genomes and their replication presumably requires a processive RNA-dependent RNA polymerase (RdRp) and enzymatic functions that suppress the consequences of the typically high error rate of viral RdRps. The arteriviruses have significantly smaller genomes and form an intriguing package with the coronaviruses to analyse viral RdRp evolution and function. The RdRp domain of nidoviruses resides in a cleavage product of the replicase polyprotein named non-structural protein (nsp) 12 in coronaviruses and nsp9 in arteriviruses. In all nidoviruses, the C-terminal RdRp domain is linked to a conserved N-terminal domain, which has been coined NiRAN (nidovirus RdRp-associated nucleotidyl transferase). Although no structural information is available, the functional characterization of the nidovirus RdRp and the larger enzyme complex of which it is part, has progressed significantly over the past decade. In coronaviruses several smaller, non-enzymatic nsps were characterized that direct RdRp function, while a 3'-to-5' exoribonuclease activity in nsp14 was implicated in fidelity. In arteriviruses, the nsp1 subunit was found to maintain the balance between genome replication and subgenomic mRNA production. Understanding RdRp behaviour and interactions during RNA synthesis and subsequent processing will be key to rationalising the evolutionary success of nidoviruses and the development of antiviral strategies.

Mapping the Nonstructural Protein Interaction Network of Porcine Reproductive and Respiratory Syndrome Virus

“…This result indicates that 1B can exert an effect on the HEL domain to promote the interaction, which is also consistent with the finding that EAV nsp10 1B interacts with the HEL domain (37). For nsp9, it is expected that there is cross talk between the NiRAN domain and RdRp (63). We actually have some evidence to support this possibility (unpublished data).…”

Identification of Nonstructural Protein 8 as the N-Terminus of the RNA-Dependent RNA Polymerase of Porcine Reproductive and Respiratory Syndrome Virus

“…The PRRSV nsp9 encodes the function of viral RdRp and is the engine driving viral RNA synthesis Beerens et al 2007;Posthuma et al 2017); it is encoded within ORF1b and translated through the C-terminal extension of nsp8 during infection (Meulenberg et al 1993;Kappes and Faaberg 2015). However, it is not clear whether the mature form of nsp9 retains nsp8, although the nsp8-9 does not contain the predictable cleavage sites for nsp2 or nsp4.…”

“…The nsp9 replicase protein is a critical determinant for the fatal virulence of the Chinese HP-PRRSV (Li et al 2014;Xu et al 2018;Zhao et al 2018). This protein specifies the putative function of viral RNA-dependent RNA polymerase (RdRp) and is the major engine for catalyzing the viral RNA synthesis during PRRSV infection Beerens et al 2007;Posthuma et al 2017). Comparative bioinformatics analyses based on the studies of equine arteritis virus (EAV) and other positive-stranded RNA viruses have localized the viral RdRp domain to the C-terminal half of nsp9 (Gorbalenya et al 1989) and assigned nidovirus RdRp-associated nucleotidyltransferase domain (NiRAN) to the N-terminal region of ORF1b-encoded portion (Lehmann et al 2015).…”

“…Comparative bioinformatics analyses based on the studies of equine arteritis virus (EAV) and other positive-stranded RNA viruses have localized the viral RdRp domain to the C-terminal half of nsp9 (Gorbalenya et al 1989) and assigned nidovirus RdRp-associated nucleotidyltransferase domain (NiRAN) to the N-terminal region of ORF1b-encoded portion (Lehmann et al 2015). As a result of the frameshift strategy, nsp9, the first protein within ORF1b, is expressed during infection as the C-terminal extension of nsp8, a small viral nsp that is encoded by ORF1a (Fang and Snijder 2010;Posthuma et al 2017). However, it has remained unclear whether the mature form of nsp9 in virus-infected cells still retains nsp8.…”

Templated Self‐Replication in Biomimetic Systems

“…[125][126][127][128] This is important as ssRNA replicases do not possess sufficient helicase activity to unwind longer double stranded RNA (dsRNA) duplex structures. [130] Notably, a Nidovirus with a genome larger than 40 kb was recently described by Saberi et al, suggesting that maintenance of large RNA genomes by these mechanisms might be possible. Thus, close coordination is required to avoid steric clashes between the replicase and translating ribosomes.…”