For most dsRNA viruses, the genome-enclosing capsid comprises 120 copies of a single capsid protein (CP) organized into 60 icosahedrally equivalent dimers, generally identified as 2 ...nonsymmetricallyinteracting CP molecules with extensive lateral contacts. The crystal structure of a partitivirus, Penicillium stoloniferum virus F (PsV-F), reveals a different organization, in which the CP dimer is related by almost-perfect local 2-fold symmetry, forms prominent surface arches, and includes extensive structure swapping between the 2 subunits. An electron cryomicroscopy map of PsV-F shows that the disordered N terminus of each CP molecule interacts with the dsRNA genome and probably participates in its packaging or transcription. Intact PsV-F particles mediate semiconservative transcription, and transcripts are likely to exit through negatively charged channels at the icosahedral 5-fold axes. Other findings suggest that the PsV-F capsid is assembled from dimers of CP dimers, with an arrangement similar to flavivirus E glycoproteins.
The complete nucleotide sequence and genome organization of a hypovirus from the isolate ME711 of
Phomopsis longicolla
was determined and compared to sequences of members of the family
Hypoviridae
. ...The genome of the hypovirus, tentatively named Phomopsis longicolla hypovirus 1 (PlHV1-ME711), was determined to be 9760 nucleotides long, excluding the 3’ poly (A) tail. The genome contains a single large open reading frame (ORF) encoding a polyprotein designated as P307. Its genomic organization is typical of members of the proposed genus
Betahypovirus
(Yaegashi et al. in Virus Res 165:143–50,
2012
).
The enigma of the transmissible disease of Helminthosporium victoriae has almost been resolved. Diseased isolates are doubly infected with two distinct viruses, the victorivirus Helminthosporium ...victoriae virus 190S and the chrysovirus HvV145S. Mixed infection, however, is not required for disease development. DNA transformation experiments and transfection assays using purified HvV190S virions strongly indicate that HvV190S alone is necessary for inducing disease symptoms. HvV145, like other chrysoviruses, appears to have no effect on colony morphology. This chapter will discuss the molecular biology of the two viruses and summarize recent results of characterization of host gene products upregulated by virus infection. Furthermore, the novel structural features of HvV190S capsid will be highlighted.
Full-length infectious cDNA clones were constructed from the genomic RNAs of three distinct strains (K-G7, K-Ha1 and K-Ho1) of the comovirus
Bean pod mottle virus (BPMV). Whereas K-G7, a subgroup I ...strain, and K-Ha1, a subgroup II strain produce mild mottling, the reassortant strain K-Ho1 (RNA1
I + RNA2
II) induces necrotic primary lesions on inoculated leaves of soybean and severe systemic leaf mottling and blistering. Pseudorecombinants of all possible combinations of transcripts were generated and tested for symptom production. Only soybean plants inoculated with combinations having RNA1 derived from the severe strain K-Ho1, regardless of the origin of RNA2, induced severe symptoms, indicating that symptom severity maps to RNA1. Experiments with chimeric RNA1 constructs indicated that the coding regions of the protease co-factor (Co-pro) and the C-terminal half of the putative helicase (Hel) are determinants of symptom severity. Symptom severity correlated well with higher accumulation of viral RNA, but neither the Co-pro nor Hel protein could be demonstrated as a suppressor of RNA silencing.
Most double-stranded RNA (dsRNA) viruses are transcribed and replicated in a specialized icosahedral capsid with a T=1 lattice consisting of 60 asymmetric capsid protein (CP) dimers. These capsids ...help to organize the viral genome and replicative complex(es). They also act as molecular sieves that isolate the virus genome from host defense mechanisms and allow the passage of nucleotides and viral transcripts. Rosellinia necatrix quadrivirus 1 (RnQV1), the type species of the family Quadriviridae, is a dsRNA fungal virus with a multipartite genome consisting of four monocistronic segments (segments 1 to 4). dsRNA-2 and dsRNA-4 encode two CPs (P2 and P4, respectively), which coassemble into ∼450-Å-diameter capsids. We used three-dimensional cryo-electron microscopy combined with complementary biophysical techniques to determine the structures of RnQV1 virion strains W1075 and W1118. RnQV1 has a quadripartite genome, and the capsid is based on a single-shelled T=1 lattice built of P2-P4 dimers. Whereas the RnQV1-W1118 capsid is built of full-length CP, P2 and P4 of RnQV1-W1075 are cleaved into several polypeptides, maintaining the capsid structural organization. RnQV1 heterodimers have a quaternary organization similar to that of homodimers of reoviruses and other dsRNA mycoviruses. The RnQV1 capsid is the first T=1 capsid with a heterodimer as an asymmetric unit reported to date and follows the architectural principle for dsRNA viruses that a 120-subunit capsid is a conserved assembly that supports dsRNA replication and organization.
Given their importance to health, members of the family Reoviridae are the basis of most structural and functional studies and provide much of our knowledge of dsRNA viruses. Analysis of bacterial, protozoal, and fungal dsRNA viruses has improved our understanding of their structure, function, and evolution, as well. Here, we studied a dsRNA virus that infects the fungus Rosellinia necatrix, an ascomycete that is pathogenic to a wide range of plants. Using three-dimensional cryo-electron microscopy and analytical ultracentrifugation analysis, we determined the structure and stoichiometry of Rosellinia necatrix quadrivirus 1 (RnQV1). The RnQV1 capsid is a T=1 capsid with 60 heterodimers as the asymmetric units. The large amount of genetic information used by RnQV1 to construct a simple T=1 capsid is probably related to the numerous virus-host and virus-virus interactions that it must face in its life cycle, which lacks an extracellular phase.
Infectious myonecrosis virus (IMNV) is an emerging pathogen of penaeid shrimp in global aquaculture. Tentatively assigned to family Totiviridae, it has a nonsegmented dsRNA genome of 7,560 bp and an ...isometric capsid of the 901-aa major capsid protein. We used electron cryomicroscopy and 3D image reconstruction to examine the IMNV virion at 8.0-Å resolution. Results reveal a totivirus-like, 120-subunit T = 1 capsid, 450 Å in diameter, but with fiber complexes protruding a further 80 Å at the fivefold axes. These protrusions likely mediate roles in the extracellular transmission and pathogenesis of IMNV, capabilities not shared by most other totiviruses. The IMNV structure is also notable in that the genome is centrally organized in five or six concentric shells. Within each of these shells, the densities alternate between a dodecahedral frame that connects the threefold axes vs. concentration around the fivefold axes, implying certain regularities in the RNA packing scheme.
Filamentous fungus Fusarium poae is a worldwide cause of the economically important disease Fusarium head blight of cereal grains. The fungus is itself commonly infected with a bisegmented dsRNA ...virus from the family Partitiviridae. For this study, we determined the structure of partitivirus Fusarium poae virus 1 (FpV1) to a resolution of 5.6Å or better by electron cryomicroscopy and three-dimensional image reconstruction. The main structural features of FpV1 are consistent with those of two other fungal partitiviruses for which high-resolution structures have been recently reported. These shared features include a 120-subunit T=1 capsid comprising 60 quasisymmetrical capsid protein dimers with both shell and protruding domains. Distinguishing features are evident throughout the FpV1 capsid, however, consistent with its more massive subunits and its greater phylogenetic divergence relative to the other two structurally characterized partitiviruses. These results broaden our understanding of conserved and variable elements of fungal partitivirus structure, as well as that of vertebrate picobirnavirus, and support the suggestion that a phylogenetic subcluster of partitiviruses closely related to FpV1 should constitute a separate taxonomic genus.
Common bean (Phaseolus vulgaris L.) is a crop of economic and nutritious importance in many parts of the world. The lack of genomic resources have impeded the advancement of common bean genomics and ...thereby crop improvement. Although concerted efforts from the "Phaseomics" consortium have resulted in the development of several genomic resources, functional studies have continued to lag due to the recalcitrance of this crop for genetic transformation.
Here we describe the use of a bean pod mottle virus (BPMV)-based vector for silencing of endogenous genes in common bean as well as for protein expression. This BPMV-based vector was originally developed for use in soybean. It has been successfully employed for both protein expression and gene silencing in this species. We tested this vector for applications in common bean by targeting common bean genes encoding nodulin 22 and stearoyl-acyl carrier protein desaturase for silencing. Our results indicate that the BPMV vector can indeed be employed for reverse genetics studies of diverse biological processes in common bean. We also used the BPMV-based vector for expressing the green fluorescent protein (GFP) in common bean and demonstrate stable GFP expression in all common bean tissues where BPMV was detected.
The availability of this vector is an important advance for the common bean research community not only because it provides a rapid means for functional studies in common bean, but also because it does so without generating genetically modified plants. Here we describe the detailed methodology and provide essential guidelines for the use of this vector for both gene silencing and protein expression in common bean. The entire VIGS procedure can be completed in 4-5 weeks.
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