Guanidine-selected mutants of poliovirus: mapping of point mutations to polypeptide 2C

Pincus, Stephanie H.; Diamond, David C.; Emini, Emilio A.; Wimmer, Eckard

doi:10.1128/jvi.57.2.638-646.1986

Cited by 167 publications

(91 citation statements)

References 51 publications

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“…GuHCl inhibits the NTPase and RNA helix-unwinding activities of NV NS3. Previous studies have reported that the NTPase and RNA helicase activities of enterovirus 2C ATPase can be inhibited by GuHCl (22,(41)(42)(43)(44)(45), which also inhibits replication of the vRNAs of a number of enteroviruses (46,47). Given the similarity between the consensus motifs in enterovirus 2C ATPase and norovirus NS3, we wanted to examine whether NV NS3 could also be a target for guanidine inhibition.…”

Section: Resultsmentioning

confidence: 99%

“…Virus-encoded RNA-remodeling proteins, particularly helicases, have long been considered valuable targets for antiviral compounds, because they are well conserved within or even across virus families (57). In particular, enterovirus 2C ATPase , another SF3 viral RNA helicase, has been found to be inhibited by ϳ1 mM GuHCl, as is enterovirus replication (22,(42)(43)(44)(45). Using an NV replicon as a model, our studies demonstrated that ϳ1 mM GuHCl can significantly inhibit the replication of HuNoV in cultured human cells with negligible cell toxicity ( Fig.…”

Section: Discussionmentioning

confidence: 99%

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Human Norovirus NS3 Has RNA Helicase and Chaperoning Activities

Hosmillo

Schwanke

et al. 2018

J Virol

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RNA-remodeling proteins, including RNA helicases and chaperones, act to remodel RNA structures and/or protein-RNA interactions and are required for all processes involving RNAs. Although many viruses encode RNA helicases and chaperones, their activities and their roles in infected cells largely remain elusive. Noroviruses are a diverse group of positive-strand RNA viruses in the family and constitute a significant and potentially fatal threat to human health. Here, we report that the protein NS3 encoded by human norovirus has both ATP-dependent RNA helicase activity that unwinds RNA helices and ATP-independent RNA-chaperoning activity that can remodel structured RNAs and facilitate strand annealing. Moreover, NS3 can facilitate viral RNA synthesis by norovirus polymerase. NS3 may therefore play an important role in norovirus RNA replication. Lastly, we demonstrate that the RNA-remodeling activity of NS3 is inhibited by guanidine hydrochloride, an FDA-approved compound, and, more importantly, that it reduces the replication of the norovirus replicon in cultured human cells. Altogether, these findings are the first to demonstrate the presence of RNA-remodeling activities encoded by and highlight the functional significance of NS3 in the noroviral life cycle. Noroviruses are a diverse group of positive-strand RNA viruses, which annually cause hundreds of millions of human infections and over 200,000 deaths worldwide. For RNA viruses, cellular or virus-encoded RNA helicases and/or chaperones have long been considered to play pivotal roles in viral life cycles. However, neither RNA helicase nor chaperoning activity has been demonstrated to be associated with any norovirus-encoded proteins, and it is also unknown whether norovirus replication requires the participation of any viral or cellular RNA helicases/chaperones. We found that a norovirus protein, NS3, not only has ATP-dependent helicase activity, but also acts as an ATP-independent RNA chaperone. Also, NS3 can facilitate viral RNA synthesis, suggesting the important role of NS3 in norovirus replication. Moreover, NS3 activities can be inhibited by an FDA-approved compound, which also suppresses norovirus replicon replication in human cells, raising the possibility that NS3 could be a target for antinoroviral drug development.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Human Norovirus NS3 Has RNA Helicase and Chaperoning Activities

Hosmillo

Schwanke

et al. 2018

J Virol

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“…This hypothesis is in agreement with the available experimental data implicating these proteins in viral RNA replication and with the reported NTP-binding capacity of TMV p126 (Evans et al 1985), although direct testing is certainly warranted. In fact, there is experimental evidence that clearly, though indirectly, demonstrates the importance of the NTP-motif in viral RNA replication, Recently several poliovirus mutants resistant to or dependent on guanidine, a potent inhibitor of RNA replication of some picornaviruses, have been thoroughly studied (Pincus et al 1986(Pincus et al , 1987. They all mapped to the 2C protein, with the amino acid replacements located in the proximity of the "A" and "B" sites of the NTPmotif, or near the conserved Asn residue in the 183rd position of the segments of 2C aligned in this paper (Fig.…”

Section: Discussionmentioning

confidence: 99%

An NTP-binding motif is the most conserved sequence in a highly diverged monophyletic group of proteins involved in positive strand RNA viral replication

et al. 1989

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NTP-motif, a consensus sequence previously shown to be characteristic of numerous NTP-utilizing enzymes, was identified in nonstructural proteins of several groups of positive-strand RNA viruses. These groups include picorna-, alpha-, and coronaviruses infecting animals and como-, poty-, tobamo-, tricorna-, hordei-, and furoviruses of plants, totalling 21 viruses. It has been demonstrated that the viral NTP-motif-containing proteins constitute three distinct families, the sequences within each family being similar to each other at a statistically highly significant level. A lower, but still valid similarity has also been revealed between the families. An overall alignment has been generated, which includes several highly conserved sequence stretches. The two most prominent of the latter contain the socalled "A" and "B" sites of the NTP-motif, with four of the five invariant amino acid residues observed within these sequences. These observations, taken together with the results of comparative analysis of the positions occupied by respective proteins (domains) in viral multidomain proteins, suggest that all the NTP-motif-containing proteins of positive-strand RNA viruses are homologous, constituting a highly diverged monophyletic group. In this group the "A" and "B" sites of the NTP-motif are the most conserved sequences and, by inference, should play the principal role in the functioning of the proteins. A hypothesis is proposed that all these proteins possess NTP-binding capacity and possibly NTPase activity, performing some NTP-dependent function in viral RNA replication. The importance of phylogenetic analysis for the assessment of the significance of the occurrence of the NTP-motif (and of sequence motifs of this sort in general) in proteins is emphasized.

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“…The above-mentioned guanidine-resistant (g r ) poliovirus mutants are a typical example. The drug targets the viral multifunctional protein 2C (414)(415)(416)(417)(418), which possesses an RNA-dependent NTPase activity (419,420) and, in particular, is hypothesized (though not proved) to function as an RNA helicase (421)(422)(423). Both g r and guanidine-dependent (g d ) classes of mutants display a wide range of phenotypes with different levels of dependence of reproductive efficiency on the guanidine concentration (416,424).…”

Section: Development Of Resistance To Mutagenic and Some Other Inhibimentioning

confidence: 99%

Emergency Services of Viral RNAs: Repair and Remodeling

Agol

Gmyl

2018

Microbiol Mol Biol Rev

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Reproduction of RNA viruses is typically error-prone due to the infidelity of their replicative machinery and the usual lack of proofreading mechanisms. The error rates may be close to those that kill the virus. Consequently, populations of RNA viruses are represented by heterogeneous sets of genomes with various levels of fitness. This is especially consequential when viruses encounter various bottlenecks and new infections are initiated by a single or few deviating genomes. Nevertheless, RNA viruses are able to maintain their identity by conservation of major functional elements. This conservatism stems from genetic robustness or mutational tolerance, which is largely due to the functional degeneracy of many protein and RNA elements as well as to negative selection. Another relevant mechanism is the capacity to restore fitness after genetic damages, also based on replicative infidelity. Conversely, error-prone replication is a major tool that ensures viral evolvability. The potential for changes in debilitated genomes is much higher in small populations, because in the absence of stronger competitors low-fit genomes have a choice of various trajectories to wander along fitness landscapes. Thus, low-fit populations are inherently unstable, and it may be said that to run ahead it is useful to stumble. In this report, focusing on picornaviruses and also considering data from other RNA viruses, we review the biological relevance and mechanisms of various alterations of viral RNA genomes as well as pathways and mechanisms of rehabilitation after loss of fitness. The relationships among mutational robustness, resilience, and evolvability of viral RNA genomes are discussed.

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Guanidine-selected mutants of poliovirus: mapping of point mutations to polypeptide 2C

Cited by 167 publications

References 51 publications

Human Norovirus NS3 Has RNA Helicase and Chaperoning Activities

Human Norovirus NS3 Has RNA Helicase and Chaperoning Activities

An NTP-binding motif is the most conserved sequence in a highly diverged monophyletic group of proteins involved in positive strand RNA viral replication

Emergency Services of Viral RNAs: Repair and Remodeling

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