Regulation of Sindbis virus RNA replication: uncleaved P123 and nsP4 function in minus-strand RNA synthesis, whereas cleaved products from P123 are required for efficient plus-strand RNA synthesis

Nonstructural proteins of Sindbis virus, nsPl, nsP2, nsP3, and nsP4, as well as intermediate polyproteins, are produced from two precursor polyproteins, Pl23 and Pl234, by a proteolytic enzyme encoded in the C-terminal half of nsP2. We studied the requirements for and the functions of the intermediate and mature processing products for Sindbis virus RNA synthesis by using site-directed mutants which have a defect(s) in processing the 1/2, 2/3, or 3/4 cleavage sites either singly or in various combinations. A mutant defective in cleaving both the 1/2 and 2/3 sites, which makes only uncleavable Pl23 and mature nsP4 as final products, produced 10-3 as much virus as did the wild-type virus after 10 hat 30°C and was nonviable at 40°C. A mutant defective in processing the 2/3 site, which makes nsPl, nsP4, and P23 as well as precursor Pl23, grew 10-1 as efficiently as wild-type virus at 30°C and 10-3 as efficiently at 40°C. Early minus-strand RNA synthesis by these mutants was as efficient as that by wild-type virus, whereas plus-strand RNA synthesis was substantially decreased compared with that by wild-type virus. A mutant defective in processing the 3/4 site was nonviable at either 30 or 40°C. The 3/4 site mutant could be complemented by the mutant unable to cleave either the 1/2 or 2/3 site, which can provide mature nsP4. We interpret these results to signify that (i) mature nsP4 is required for RNA replication, (ii) nsP4 and uncleaved Pl23 function in minus-strand RNA synthesis, and (iii) cleavage of Pl23 is required for efficient plus-strand RNA synthesis. We propose that Sindbis virus RNA replication is regulated by differential proteolysis of Pl23. Early in infection, nsP4 and uncleaved Pl23 form transient minus-strand RNA replication complexes which vanish upon cleavage of Pl23. Later in infection, an elevated level of viral proteinase activity eliminates de novo synthesis of Pl23, and no further synthesis of minus-strand RNA is possible. In contrast, nsP4 and cleavage products from Pl23 form plus-strand RNA replication complexes which are stable and remain active throughout the infection cycle

Taxonomy of the order Bunyavirales : second update 2018

In October 2018, the order Bunyavirales was amended by inclusion of the family Arenaviridae, abolishment of three families, creation of three new families, 19 new genera, and 14 new species, and renaming of three genera and 22 species. This article presents the updated taxonomy of the order Bunyavirales as now accepted by the International Committee on Taxonomy of Viruses (ICTV).Non peer reviewe

Taxonomy of the family Arenaviridae and the order Bunyavirales : update 2018

In 2018, the family Arenaviridae was expanded by inclusion of 1 new genus and 5 novel species. At the same time, the recently established order Bunyavirales was expanded by 3 species. This article presents the updated taxonomy of the family Arenaviridae and the order Bunyavirales as now accepted by the International Committee on Taxonomy of Viruses (ICTV) and summarizes additional taxonomic proposals that may affect the order in the near future.Peer reviewe

2021 Taxonomic update of phylum Negarnaviricota (Riboviria: Orthornavirae), including the large orders Bunyavirales and Mononegavirales.

Correction to: 2021 Taxonomic update of phylum Negarnaviricota (Riboviria: Orthornavirae), including the large orders Bunyavirales and Mononegavirales. Archives of Virology (2021) 166:3567–3579. https://doi.org/10.1007/s00705-021-05266-wIn March 2021, following the annual International Committee on Taxonomy of Viruses (ICTV) ratification vote on newly proposed taxa, the phylum Negarnaviricota was amended and emended. The phylum was expanded by four families (Aliusviridae, Crepuscuviridae, Myriaviridae, and Natareviridae), three subfamilies (Alpharhabdovirinae, Betarhabdovirinae, and Gammarhabdovirinae), 42 genera, and 200 species. Thirty-nine species were renamed and/or moved and seven species were abolished. This article presents the updated taxonomy of Negarnaviricota as now accepted by the ICTV.This work was supported in part through Laulima Government Solutions, LLC prime contract with the US National Institute of Allergy and Infectious Diseases (NIAID) under Contract No. HHSN272201800013C. J.H.K. performed this work as an employee of Tunnell Government Services (TGS), a subcontractor of Laulima Government Solutions, LLC under Contract No. HHSN272201800013C. This work was also supported in part with federal funds from the National Cancer Institute (NCI), National Institutes of Health (NIH), under Contract No. 75N91019D00024, Task Order No. 75N91019F00130 to I.C., who was supported by the Clinical Monitoring Research Program Directorate, Frederick National Lab for Cancer Research. This work was also funded in part by Contract No. HSHQDC-15-C-00064 awarded by DHS S&T for the management and operation of The National Biodefense Analysis and Countermeasures Center, a federally funded research and development center operated by the Battelle National Biodefense Institute (V.W.); and NIH contract HHSN272201000040I/HHSN27200004/D04 and grant R24AI120942 (N.V., R.B.T.). S.S. acknowledges partial support from the Special Research Initiative of Mississippi Agricultural and Forestry Experiment Station (MAFES), Mississippi State University, and the National Institute of Food and Agriculture, US Department of Agriculture, Hatch Project 1021494. Part of this work was supported by the Francis Crick Institute which receives its core funding from Cancer Research UK (FC001030), the UK Medical Research Council (FC001030), and the Wellcome Trust (FC001030).S

2020 taxonomic update for phylum Negarnaviricota (Riboviria: Orthornavirae), including the large orders Bunyavirales and Mononegavirales.

In March 2020, following the annual International Committee on Taxonomy of Viruses (ICTV) ratification vote on newly proposed taxa, the phylum Negarnaviricota was amended and emended. At the genus rank, 20 new genera were added, two were deleted, one was moved, and three were renamed. At the species rank, 160 species were added, four were deleted, ten were moved and renamed, and 30 species were renamed. This article presents the updated taxonomy of Negarnaviricota as now accepted by the ICTV

2021 Taxonomic update of phylum Negarnaviricota (Riboviria: Orthornavirae), including the large orders Bunyavirales and Mononegavirales.

In March 2021, following the annual International Committee on Taxonomy of Viruses (ICTV) ratification vote on newly proposed taxa, the phylum Negarnaviricota was amended and emended. The phylum was expanded by four families (Aliusviridae, Crepuscuviridae, Myriaviridae, and Natareviridae), three subfamilies (Alpharhabdovirinae, Betarhabdovirinae, and Gammarhabdovirinae), 42 genera, and 200 species. Thirty-nine species were renamed and/or moved and seven species were abolished. This article presents the updated taxonomy of Negarnaviricota as now accepted by the ICTV

Non-AUG Translation Initiation in a Plant RNA Virus: a Forty-Amino-Acid Extension Is Added to the N Terminus of the Soil-Borne Wheat Mosaic Virus Capsid Protein

RNA 2 of soil-borne wheat mosaic virus (SBWMV), the type species of the genus Furovirus, encodes a protein previously hypothesized to be initiated at an in-frame non-AUG codon upstream of the AUG initiation codon (nucleotide positions 334 to 336) for the 19-kDa capsid protein. Site-directed mutagenesis and in vitro transcription and translation analysis indicated that CUG (nucleotides 214 to 216) is the initiation codon for a protein with a calculated molecular mass of 25 kDa composed of a 40-amino-acid extension to the N terminus of the 19-kDa capsid protein. A stable deletion mutant, which was isolated after extensive passages of a wild-type SBWMV, contained a mixture of two deleted RNA 2’s, only one of which coded for the 25-kDa protein. The amino acid sequence of the N-terminal extension was moderately conserved and the CUG initiation codon was preserved among three SBWMV isolates from Japan and the United States. This amino acid sequence conservation, as well as the retention of expression of the 25-kDa protein in the stable deletion mutant, suggests that the 25-kDa protein is functional in the life cycle of SBWMV. This is the first report of a non-AUG translation initiation in a plant RNA virus genome

SOIL-BORNE WHEAT MOSAIC VIRUS HAS TWO GENOMIC RNA COMPONENTS, THE SMALLER OF WHICH UNDERGOES FREQUENT DELETION MUTATION

Soil-borne wheat mosaic virus (SBWMV) has rod-shaped virions 20nm in diameter and lengths of 281 and 138nm, but in some isolates a 92nm particle is also present. Successive manual transfer of the latter isolate had given virus with 281 and 92nm long virions. This study concerns function and relationships of the three components. RNAs of 281, 138, and 92nm long virions (designated here by relative lengths as 1.0L, 0.5L, and 0.35L, respectively) were purified by three cycles of sucrose density gradient centrifugation. Infectivity assay with these RNAs proved the bipartite nature of SBWMV, the combination of 1.0L and either 0.5L or 0.35L RNAs being required for infection and for multiplication of progeny viruses. Infection of plants with 1.0L and 0.35L RNAs gave progeny of 1.0L and 0.35L only, but infection with 1.0L and 0.5L RNAs gave progeny with 1.0L and 0.5L, and also variants with various sizes smaller than 0.5L. Virions of 0.4L and 0.35L were functional in combination with 1.0L virion. Similarly, infection of plants by manual inoculation with 1.0L and 0.5L virions resulted in variants smaller than 0.5L. The ratio of the smaller variants to 0.5L virions increased with time. Variants smaller than 0.5L were found in wheat plants infected by the vector fungus, Polymyxa graminis, transplanted from the field in November and grown at 17(DEGREES) in a greenhouse until March, whereas virus from plants taken from the same field in March had 1.0L and 0.5L virions only. Regardless of the method of inoculation, the size profile of the variants varied from plant to plant. These results indicate that 0.5L RNA and SBWMV is unstable and produces deletion mutants when the host plants are grown at 17(DEGREES), with 0.35L RNA as the smallest functional molecule. The reason why 1.0L and 0.5L virions dominate the virion species infecting wheat plants in early spring is uncertain. The coat proteins of all isolates had Mr of 19,700. The Mr\u27s of RNAs of 1.0L, 0.5L, 0.4L, and 0.35L virions, determined under denaturing conditions, were 2.28, 1.23, 0.97, and 0.86 x 10(\u276), respectively. A new virus group, furovirus (fungus-borne rod-shaped virus) is proposed for SBWMV

Two Purified RNAs of Soil-borne Wheat Mosaic Virus are Needed for Infection

RNAs of soil-borne wheat mosaic virus (SBWMV) from virions 281 nm, 138 nm and 92 nm long (designated here by relative lengths as 1∙0L, 0∙SL and 0∙35L, respectively), were isolated and purified by three cycles of sucrose density gradient centrifugation. Infectivity assays with these RNAs proved the bipartite nature of SBWMV, the combination of 1∙0L and either 0∙5L or 0∙35L RNAs being required for infection and for multiplication of progeny viruses. The 0∙5L RNA underwent deletion mutation, producing smaller variants with various sizes, of which 0∙4L and 0∙35L RNAs were confirmed to be functional in combination with 1∙0L RNA. The coat proteins of all isolates had mol. wt. of 19700. The mol. wt. of 1∙0L, 0∙5L, 0∙4L and 0∙35L RNAs, determined under denaturing conditions, were 2∙28 × 106 (6500 bases), 1∙23 x 106 (3500 bases), 0-97 x 106 (2800 bases) and 0∙86 x 106 (2450 bases), respectively. A new virus group, furovirus (fungus-borne rod-shaped virus), is proposed for SBWMV