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The nonstructural protein 2C of Coxsackie B virus has RNA helicase and chaperoning activities.

ABSTRACT: RNA-remodeling proteins, including RNA helicases and chaperones, play vital roles in the remodeling of structured RNAs. During viral replication, viruses require RNA-remodeling proteins to facilitate proper folding and/or re-folding the viral RNA elements. Coxsackieviruses B3 (CVB3) and Coxsackieviruses B5 (CVB5), belonging to the genus Enterovirus in the family Picornaviridae, have been reported to cause various infectious diseases such as hand-foot-and-mouth disease, aseptic meningitis, and viral myocarditis. However, little is known about whether CVB3 and CVB5 encode any RNA remodeling proteins. In this study, we showed that 2C proteins of CVB3 and CVB5 contained the conserved SF3 helicase A, B, and C motifs, and functioned not only as RNA helicase that unwound RNA helix bidirectionally in an NTP-dependent manner, but also as RNA chaperone that remodeled structured RNAs and facilitated RNA strand annealing independently of NTP. In addition, we determined that the NTPase activity and RNA helicase activity of 2C proteins of CVB3 and CVB5 were dependent on the presence of divalent metallic ions. Our findings demonstrate that 2C proteins of CVBs possess RNA-remodeling activity and underline the functional importance of 2C protein in the life cycle of CVBs.

SUBMITTER: Chen Z

PROVIDER: S-EPMC9583185 | biostudies-literature | 2022 Oct

REPOSITORIES: biostudies-literature

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The nonstructural protein 2C of Coxsackie B virus has RNA helicase and chaperoning activities.

Chen Ziyu Z Xiong Xiaobei X Li Yiyang Y Huang Muhan M Ren Yujie Y Wu Di D Qiu Yang Y Chen Mingzhou M Shu Ting T Zhou Xi X

Virologica Sinica 20220516 5

RNA-remodeling proteins, including RNA helicases and chaperones, play vital roles in the remodeling of structured RNAs. During viral replication, viruses require RNA-remodeling proteins to facilitate proper folding and/or re-folding the viral RNA elements. Coxsackieviruses B3 (CVB3) and Coxsackieviruses B5 (CVB5), belonging to the genus Enterovirus in the family Picornaviridae, have been reported to cause various infectious diseases such as hand-foot-and-mouth disease, aseptic meningitis, and vi ...[more]

PMID: 35589079

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Project description: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 in vitro activities and their roles in infected cells largely remain elusive. Noroviruses are a diverse group of positive-strand RNA viruses in the family Caliciviridae 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 in vitro 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 Caliciviridae and highlight the functional significance of NS3 in the noroviral life cycle.IMPORTANCE 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 in vitro 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.

Project description:Enteroviruses belong to the genus Enterovirus of the family Picornaviridae and include four human enterovirus groups (EV-A to -D): the epidemic of enteroviruses such as human enterovirus A71 (EV-A71) and coxsackievirus A16 (CVA16) is a threat to global public health. Enteroviral protein 2C is the most conserved nonstructural protein among all enteroviruses and possesses RNA helicase activity that plays pivotal roles during enteroviral life cycles, which makes 2C an attractive target for developing antienterovirus drugs. In this study, we designed a peptide, named 2CL, based on the structure of EV-A71 2C. This peptide effectively impaired the oligomerization of EV-A71 2C protein and inhibited the RNA helicase activities of 2C proteins encoded by EV-A71 and CVA16, both of which belong to EV-A, and showed potent antiviral efficacy against EV-A71 and CVA16 in cells. Moreover, the 2CL treatment elicited a strong in vivo protective efficacy against lethal EV-A71 challenge. In addition, the antiviral strategy of targeting the 2C helicase activity can be applied to inhibit the replication of EV-B. Either 2CL or B-2CL, the peptide redesigned based on the 2CL-corresponding sequence of EV-Bs, could exert effective antiviral activity against two important EV-Bs, coxsackievirus B3 and echovirus 11. Together, our findings demonstrated that targeting the helicase activity of 2C with a rationally designed peptide is an efficient antiviral strategy against enteroviruses, and 2CL and B-2CL show promising clinical potential to be further developed as broad-spectrum antienterovirus drugs.IMPORTANCE Enteroviruses are a large group of positive-sense single-stranded RNA viruses and include numerous human pathogens, such as enterovirus A71 (EV-A71), coxsackieviruses, and echoviruses. However, no approved EV antiviral drugs are available. Enteroviral 2C is the most conserved nonstructural protein among all enteroviruses and contains the RNA helicase activity critical for the viral life cycle. Herein, according to the structure of EV-A71 2C, we designed a peptide that effectively inhibited the RNA helicase activities of EV-A71- and coxsackievirus A16 (CVA16)-encoded 2C proteins. Moreover, this peptide exerted potent antiviral effects against EV-A71 and CVA16 in cells and elicited therapeutic efficacy against lethal EV-A71 challenge in vivo Furthermore, we demonstrate that the strategy of targeting the 2C helicase activity can be used for other relevant enteroviruses, including coxsackievirus B3 and echovirus 11. In summary, our findings provide compelling evidence that the designed peptides targeting the helicase activity of 2C could be broad-spectrum antivirals for enteroviruses.

Project description:RNA helicases and chaperones are the two major classes of RNA remodeling proteins, which function to remodel RNA structures and/or RNA-protein interactions, and are required for all aspects of RNA metabolism. Although some virus-encoded RNA helicases/chaperones have been predicted or identified, their RNA remodeling activities in vitro and functions in the viral life cycle remain largely elusive. Enteroviruses are a large group of positive-stranded RNA viruses in the Picornaviridae family, which includes numerous important human pathogens. Herein, we report that the nonstructural protein 2CATPase of enterovirus 71 (EV71), which is the major causative pathogen of hand-foot-and-mouth disease and has been regarded as the most important neurotropic enterovirus after poliovirus eradication, functions not only as an RNA helicase that 3'-to-5' unwinds RNA helices in an adenosine triphosphate (ATP)-dependent manner, but also as an RNA chaperone that destabilizes helices bidirectionally and facilitates strand annealing and complex RNA structure formation independently of ATP. We also determined that the helicase activity is based on the EV71 2CATPase middle domain, whereas the C-terminus is indispensable for its RNA chaperoning activity. By promoting RNA template recycling, 2CATPase facilitated EV71 RNA synthesis in vitro; when 2CATPase helicase activity was impaired, EV71 RNA replication and virion production were mostly abolished in cells, indicating that 2CATPase-mediated RNA remodeling plays a critical role in the enteroviral life cycle. Furthermore, the RNA helicase and chaperoning activities of 2CATPase are also conserved in coxsackie A virus 16 (CAV16), another important enterovirus. Altogether, our findings are the first to demonstrate the RNA helicase and chaperoning activities associated with enterovirus 2CATPase, and our study provides both in vitro and cellular evidence for their potential roles during viral RNA replication. These findings increase our understanding of enteroviruses and the two types of RNA remodeling activities.

Dataset Information

The nonstructural protein 2C of Coxsackie B virus has RNA helicase and chaperoning activities.

Publications

The nonstructural protein 2C of Coxsackie B virus has RNA helicase and chaperoning activities.

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