Pepino mosaic virus capsid protein interacts with a tomato heat shock protein cognate 70

Mathioudakis, Matthaios M.; Veiga, R.; Ghita, Melania; Tsikou, Daniela; Medina, Vicente; Canto, Tomás; Makris, Antonios M.; Livieratos, Ioannis C.

doi:10.1016/j.virusres.2011.08.007

Cited by 41 publications

(44 citation statements)

References 88 publications

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“…It is also possible that heat-shocked plants support greater cell-to-cell movement of CNV, as has been reported previously for other viruses (9,22,37). In addition, heatshocked plants might display compromised resistance to CNV, allowing for increased spread.…”

Section: Resultsmentioning

confidence: 62%

“…Hsp70 is activated under stress conditions such as heat, virus infection, and oxidative stress (6,(10)(11)(12)(13)(14)(15)(16)(17)(18)(19). Whereas Hsc70 is generally expressed constitutively, it can also be induced by environmental stresses such as thermal and pesticide stress, as well as virus infection (10,(20)(21)(22)(23). It has also been found that Hsp70 and Hsc70 may complement each other in a synergistic manner to preserve cellular integrity during metabolic challenges (24).…”

mentioning

confidence: 99%

“…HSP70 homologs have also been found to be involved in the cell-to-cell movement of closteroviruses and tomato yellow leaf curl virus (37,44,45). Hsc70 has been found to interact with the coat protein and virions of pepino mosaic virus and colocalizes with virions in the phloem of infected plants (22). HSP70s have also been found to be involved in the assembly of the replicase complex of tomato bushy stunt virus (TBSV) and red clover necrotic mosaic virus (46,47) and positively regulate the replication process of several other RNA viruses, including potyvirus, pepino mosaic virus, rabies virus, rice stripe virus, tomato yellow leaf curl virus, and respiratory syncytial virus (41,(48)(49)(50)(51).…”

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confidence: 99%

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Cucumber Necrosis Virus Recruits Cellular Heat Shock Protein 70 Homologs at Several Stages of Infection

Alam

Rochon

2016

J Virol

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RNA viruses often depend on host factors for multiplication inside cells due to the constraints of their small genome size and limited coding capacity. One such factor that has been exploited by several plant and animal viruses is heat shock protein 70 (HSP70) family homologs which have been shown to play roles for different viruses in viral RNA replication, viral assembly, disassembly, and cell-to-cell movement. Using next generation sequence analysis, we reveal that several isoforms of Hsp70 and Hsc70 transcripts are induced to very high levels during cucumber necrosis virus (CNV) infection of Nicotiana benthamiana and that HSP70 proteins are also induced by at least 10-fold. We show that HSP70 family protein homologs are co-opted by CNV at several stages of infection. We have found that overexpression of Hsp70 or Hsc70 leads to enhanced CNV genomic RNA, coat protein (CP), and virion accumulation, whereas downregulation leads to a corresponding decrease. Hsc70-2 was found to increase solubility of CNV CP in vitro and to increase accumulation of CNV CP independently of viral RNA replication during coagroinfiltration in N. benthamiana. In addition, virus particle assembly into virus-like particles in CP agroinfiltrated plants was increased in the presence of Hsc70-2. HSP70 was found to increase the targeting of CNV CP to chloroplasts during infection, reinforcing the role of HSP70 in chloroplast targeting of host proteins. Hence, our findings have led to the discovery of a highly induced host factor that has been co-opted to play multiple roles during several stages of the CNV infection cycle. IMPORTANCEBecause of the small size of its RNA genome, CNV is dependent on interaction with host cellular components to successfully complete its multiplication cycle. We have found that CNV induces HSP70 family homologs to a high level during infection, possibly as a result of the host response to the high levels of CNV proteins that accumulate during infection. Moreover, we have found that CNV co-opts HSP70 family homologs to facilitate several aspects of the infection process such as viral RNA, coat protein and virus accumulation. Chloroplast targeting of the CNV CP is also facilitated, which may aid in CNV suppression of host defense responses. Several viruses have been shown to induce HSP70 during infection and others to utilize HSP70 for specific aspects of infection such as replication, assembly, and disassembly. We speculate that HSP70 may play multiple roles in the infection processes of many viruses. C ucumber necrosis virus (CNV) is a positive-strand RNA virus in the genus Tombusvirus, family Tombusviridae (1). The CNV genome is monopartite and consists of ϳ4.7 kb of positive polarity single-stranded RNA. The genome contains five open reading frames (ORFs) which encode five different proteins: the auxiliary replicase factor (p33), the RNA-dependent RNA polymerase (RdRp) (p92), the coat protein (CP; p41), the movement protein (p21), and the silencing suppressor (p20) (Fig. 1A). The UAG stop codon of the p...

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Section: Resultsmentioning

confidence: 62%

mentioning

confidence: 99%

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confidence: 99%

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Cucumber Necrosis Virus Recruits Cellular Heat Shock Protein 70 Homologs at Several Stages of Infection

Alam

Rochon

2016

J Virol

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“…In addition to protecting viral genomes by forming protective capsids, they can affect viral pathogenesis through its implication in almost all stages of the viral life cycle, beginning with processes that enhance virus entry into plant cells to those that facilitate the transmission of viruses from plant to plant. For example, PepMV CP has been shown to interact with a tomato heat shock protein and thereby influence viral symptoms (19).…”

Section: Discussionmentioning

confidence: 99%

Generation of an Attenuated, Cross-Protective Pepino mosaic virus Variant Through Alignment-Guided Mutagenesis of the Viral Capsid Protein

et al. 2015

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Mild variants of many viruses are able to protect infected plants from subsequent invasion by more severe variants of the same viruses through a process known as cross-protection. In the past, the cross-protective viral variants were commonly derived from mild field isolates that were sometimes genetically heterogeneous, providing variable levels of cross-protection. Here, we report a novel approach to rapidly generate cross-protective variants of the tomato-infecting Pepino mosaic virus (PepMV) independently of the availability of mild field isolates. Our approach sought to attenuate PepMV by mutating less conserved amino acid residues of the abundantly produced capsid protein (CP). These less-conserved amino acid residues were identified through multiple alignments of CPs of six potexviruses including PepMV, and were altered systematically to yield six PepMV mutants. These mutants were subsequently inoculated onto the model plant Nicotiana benthamiana, as well as tomato, to evaluate their accumulation levels, symptom severities, and cross-protection potentials. The mutant KD, in which the threonine (T) and alanine (A) residues at CP positions 66 and 67 were replaced with lysine (K) and aspartic acid (D), respectively, were found to accumulate to low levels in infected plants, cause very mild symptoms, and effectively protect both N. benthamiana and tomato against secondary infections by wild-type PepMV. These data suggest that our approach represents a simple, fast, and reliable way of generating attenuated viral variants capable of cross-protection.

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“…Several reports, particularly on positive-strand RNA viruses of plants, examined the key role of cellular chaperones and folding enzymes. During plant virus infection, several Hsp proteins are involved in the formation of membrane-bound RCs for certain members of the Tombusvirus, Tobamovirus, Potyvirus, and Potexvirus genera (50)(51)(52)(53). There are several cochaperones, including Hsp40, that associate with the bromovirus RC (54)(55)(56).…”

Section: Discussionmentioning

confidence: 99%

Chaperone-Assisted Protein Folding Is Critical for Yellow Fever Virus NS3/4A Cleavage and Replication

Bozzacco

Andréo

et al. 2016

J Virol

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DNAJC14, a heat shock protein 40 (Hsp40) cochaperone, assists with Hsp70-mediated protein folding. Overexpressed DNAJC14 is targeted to sites of yellow fever virus (YFV) replication complex (RC) formation, where it interacts with viral nonstructural (NS) proteins and inhibits viral RNA replication. How RCs are assembled and the roles of chaperones in this coordinated process are largely unknown. We hypothesized that chaperones are diverted from their normal cellular protein quality control function to play similar roles during viral infection. Here, we show that DNAJC14 overexpression affects YFV polyprotein processing and alters RC assembly. We monitored YFV NS2A-5 polyprotein processing by the viral NS2B-3 protease in DNAJC14-overexpressing cells. Notably, DNAJC14 mutants that did not inhibit YFV replication had minimal effects on polyprotein processing, while overexpressed wild-type DNAJC14 affected the NS3/4A and NS4A/2K cleavage sites, resulting in altered NS3-to-NS3-4A ratios. This suggests that DNAJC14's folding activity normally modulates NS3/4A/2K cleavage events to liberate appropriate levels of NS3 and NS4A and promote RC formation. We introduced amino acid substitutions at the NS3/4A site to alter the levels of the NS3 and NS4A products and examined their effects on YFV replication. Residues with reduced cleavage efficiency did not support viral RNA replication, and only revertant viruses with a restored wild-type arginine or lysine residue at the NS3/4A site were obtained. We conclude that DNAJC14 inhibition of RC formation upon DNAJC14 overexpression is likely due to chaperone dysregulation and that YFV probably utilizes DNAJC14's cochaperone function to modulate processing at the NS3/4A site as a mechanism ensuring virus replication. IMPORTANCEFlaviviruses are single-stranded RNA viruses that cause a wide range of illnesses. Upon host cell entry, the viral genome is translated on endoplasmic reticulum (ER) membranes to produce a single polyprotein, which is cleaved by host and viral proteases to generate viral proteins required for genome replication and virion production. Several studies suggest a role for molecular chaperones during these processes. While the details of chaperone roles have been elusive, in this report we show that overexpression of the ER-resident cochaperone DNAJC14 affects YFV polyprotein processing at the NS3/4A site. This work reveals that DNAJC14 modulation of NS3/4A site processing is an important mechanism to ensure virus replication. Our work highlights the importance of finely regulating flavivirus polyprotein processing. In addition, it suggests future studies to address similarities and/or differences among flaviviruses and to interrogate the precise mechanisms employed for polyprotein processing, a critical step that can ultimately be targeted for novel drug development.

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Pepino mosaic virus capsid protein interacts with a tomato heat shock protein cognate 70

Cited by 41 publications

References 88 publications

Cucumber Necrosis Virus Recruits Cellular Heat Shock Protein 70 Homologs at Several Stages of Infection

Cucumber Necrosis Virus Recruits Cellular Heat Shock Protein 70 Homologs at Several Stages of Infection

Generation of an Attenuated, Cross-Protective Pepino mosaic virus Variant Through Alignment-Guided Mutagenesis of the Viral Capsid Protein

Chaperone-Assisted Protein Folding Is Critical for Yellow Fever Virus NS3/4A Cleavage and Replication

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