Crystal Structure of the Superfamily 1 Helicase from Tomato Mosaic Virus

Nishikiori, Masaki; Sugiyama, Shigeru; Xiang, Hongyu; Niiyama, Mayumi; Ishibashi, Kazuhiro; Inoue, Tsuyoshi; Ishikawa, Masayuki; Matsumura, Hideki; Katoh, Etsuko

doi:10.1128/jvi.00118-12

Cited by 56 publications

(37 citation statements)

References 52 publications

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“…We recently determined a crystal structure for the helicase domain of ToMV 130K protein (residues 666 to 1116), which consists of a C-terminal helicase core containing two RecA folds and an N-terminal accessory domain (49). Yeast two-hybrid experiments designed in light of the structure information suggested that ARL8 interacts with the N-terminal accessory domain and TOM1 interacts with both the accessory domain and the helicase core (49).…”

Section: Discussionmentioning

confidence: 99%

“…Yeast two-hybrid experiments designed in light of the structure information suggested that ARL8 interacts with the N-terminal accessory domain and TOM1 interacts with both the accessory domain and the helicase core (49). Moreover, the reporter activity in the yeast two-hybrid experiment was much stronger between ARL8 and the accessory domain alone than between ARL8 and the full-length helicase domain polypeptide, suggesting that conformational changes in the 130K protein to expose the ARL8-binding site are associated with the process of complex formation (49). On the other hand, the mutation sites of the resistance-breaking ToMV mutants are located on the surface of the helicase core that is opposite the surface where the accessory domain is located.…”

Section: Discussionmentioning

confidence: 99%

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The Resistance Protein Tm-1 Inhibits Formation of a Tomato Mosaic Virus Replication Protein-Host Membrane Protein Complex

Ishibashi

Ishikawa

2013

J Virol

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The Tm-1 gene of tomato confers resistance to Tomato mosaic virus (ToMV). Tm-1 encodes a protein that binds ToMV replication proteins and inhibits the RNA-dependent RNA replication of ToMV. The replication proteins of resistance-breaking mutants of ToMV do not bind Tm-1, indicating that the binding is important for inhibition. In this study, we analyzed how Tm-1 inhibits ToMV RNA replication in a cell-free system using evacuolated tobacco protoplast extracts. In this system, ToMV RNA replication is catalyzed by replication proteins bound to membranes, and the RNA polymerase activity is unaffected by treatment with 0.5 M NaCl-containing buffer and remains associated with membranes. We show that in the presence of Tm-1, negative-strand RNA synthesis is inhibited; the replication proteins associate with membranes with binding that is sensitive to 0.5 M NaCl; the viral genomic RNA used as a translation template is not protected from nuclease digestion; and host membrane proteins TOM1, TOM2A, and ARL8 are not copurified with the membrane-bound 130K replication protein. Deletion of the polymerase read-through domain or of the 3= untranslated region (UTR) of the genome did not prevent the formation of complexes between the 130K protein and the host membrane proteins, the 0.5 M NaCl-resistant binding of the replication proteins to membranes, and the protection of the genomic RNA from nucleases. These results indicate that Tm-1 binds ToMV replication proteins to inhibit key events in replication complex formation on membranes that precede negative-strand RNA synthesis. Tomato mosaic virus (ToMV) is a positive-strand RNA virus in the genus Tobamovirus, family Virgaviridae (1). The genome of a tobamovirus encodes at least four proteins (2). The 130K protein and its read-through product, the 180K protein, are involved in RNA replication and thus are referred to as replication proteins (3). The 130K protein contains the methyltransferase and helicase domains, and the read-through region of the 180K protein contains the polymerase domain. The other two tobamoviral proteins, movement protein and coat protein, are not required for RNA replication (4, 5).When the genomic RNA of a positive-strand RNA virus enters host cells, the replication proteins are translated from the genomic RNA, recognize the genomic RNA as a template for replication, and form replication complexes on intracellular membranes. Replication of all known eukaryotic positive-strand RNA viruses occurs in replication complexes formed on host membranes (6-10), which contain viral genomic RNAs, replication proteins, and host factors (11). To avoid elicitation of host defense systems triggered by viral double-stranded RNA (12), the activity of eukaryotic positive-strand RNA virus RNA-dependent RNA polymerases (RdRps) must be strictly regulated so that they synthesize negative-strand RNA only in the replication complexes that are sequestered from the cytoplasm. Many host factors, including chaperones and enzymes, and cellular membranes are hijacked by viruses for rep...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

The Resistance Protein Tm-1 Inhibits Formation of a Tomato Mosaic Virus Replication Protein-Host Membrane Protein Complex

Ishibashi

Ishikawa

2013

J Virol

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“…30 Crystal structure of the helicase domain of ToMV superfamily 1 helicase solved recently has provided key insight into the helicase region required for binding to Arl8 and TOM1. 53 …”

Section: Arl8b Regulates Phagosome-lysosome Fusion and Microbial Pathmentioning

confidence: 99%

Arf-like GTPase Arl8: Moving from the periphery to the center of lysosomal biology

Khatter¹,

Sindhwani

Sharma³

2015

Cellular Logistics

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“…The amino acid changes found in Tm-1 resistance-breaking mutants locate at the helicase domain of ToMV replication proteins (ToMV-Hel) and decrease the affinity of Tm-1 for the replication proteins (12). We recently determined a crystal structure of ToMV-Hel (residues S666-Q1116 of the replication proteins) (15). ToMV-Hel contains two canonical RecA-like α/β domains (1A and 2A), which form the helicase core, and a structurally unique N-terminal domain.…”

Section: Significancementioning

confidence: 99%

“…ToMV-Hel contains two canonical RecA-like α/β domains (1A and 2A), which form the helicase core, and a structurally unique N-terminal domain. The positions of mutations in Tm-1 resistance-breaking mutants have been mapped to the surface of the molecule (15).…”

Section: Significancementioning

confidence: 99%

Structural basis for the recognition–evasion arms race between Tomato mosaic virus and the resistance gene Tm-1

Ishibashi

Kezuka

Kobayashi

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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The tomato mosaic virus (ToMV) resistance gene Tm-1 encodes a protein that shows no sequence homology to functionally characterized proteins. Tm-1 binds ToMV replication proteins and thereby inhibits replication complex formation. ToMV mutants that overcome this resistance have amino acid substitutions in the helicase domain of the replication proteins (ToMV-Hel). A small region of Tm-1 in the genome of the wild tomato Solanum habrochaites has been under positive selection during its antagonistic coevolution with ToMV. Here we report crystal structures for the N-terminal inhibitory domains of Tm-1 and a natural Tm-1 variant with an I91-to-T substitution that has a greater ability to inhibit ToMV RNA replication and their complexes with ToMV-Hel. Each complex contains a Tm-1 dimer and two ToMV-Hel monomers with the interfaces between Tm-1 and ToMV-Hel bridged by ATP. Residues in ToMV-Hel and Tm-1 involved in antagonistic coevolution are found at the interface. The structural differences between ToMV-Hel in its free form and in complex with Tm-1 suggest that Tm-1 affects nucleoside triphosphatase activity of ToMV-Hel, and this effect was confirmed experimentally. Molecular dynamics simulations of complexes formed by Tm-1 with ToMV-Hel variants showed how the amino acid changes in ToMV-Hel impair the interaction with Tm-1 to overcome the resistance. With these findings, together with the biochemical properties of the interactions between ToMV-Hel and Tm-1 variants and effects of the mutations in the polymorphic residues of Tm-1, an atomic view of a step-bystep coevolutionary arms race between a plant resistance protein and a viral protein emerges.protein complex | virus resistance

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Crystal Structure of the Superfamily 1 Helicase from Tomato Mosaic Virus

Cited by 56 publications

References 52 publications

The Resistance Protein Tm-1 Inhibits Formation of a Tomato Mosaic Virus Replication Protein-Host Membrane Protein Complex

The Resistance Protein Tm-1 Inhibits Formation of a Tomato Mosaic Virus Replication Protein-Host Membrane Protein Complex

Arf-like GTPase Arl8: Moving from the periphery to the center of lysosomal biology

Structural basis for the recognition–evasion arms race between Tomato mosaic virus and the resistance gene Tm-1

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