scholarly journals Discovery of divided RdRp sequences and a hitherto unknown genomic complexity in fungal viruses

2020 ◽  
Author(s):  
Yuto Chiba ◽  
Takashi Yaguchi ◽  
Syun-ichi Urayama ◽  
Daisuke Hagiwara
Keyword(s):  
Rna Polymerase ◽  
Rna Viruses ◽  
Current Knowledge ◽  
Single Gene ◽  
Rna Seq ◽  
Rna Dependent Rna Polymerase ◽  
Independent Manner ◽  
Viral Genomes ◽  
True Diversity ◽  
Fungal Viruses

AbstractBy identifying variations in viral RNA genomes, cutting-edge metagenome technology has potential to reshape current concepts about the evolution of RNA viruses. This technology, however, cannot process low-homology genomic regions properly, leaving the true diversity of RNA viruses unappreciated. To overcome this technological limitation we applied an advanced method, Fragmented and Primer-Ligated Double-stranded (ds) RNA Sequencing (FLDS), to screen RNA viruses from 155 fungal isolates, which allowed us to obtain complete viral genomes in a homology-independent manner. We created a high-quality catalog of 19 RNA viruses (12 viral species) that infect Aspergillus isolates. Among them, nine viruses were not detectable by the conventional methodology involving agarose gel electrophoresis of dsRNA, a hallmark of RNA virus infections. Segmented genome structures were determined in 42% of the viruses. Some RNA viruses had novel genome architectures; one contained a dual methyltransferase domain and another had a separated RNA-dependent RNA polymerase (RdRp) gene. A virus from a different fungal taxon (Pyricularia) had an RdRp sequence that was separated on different segments, suggesting that a divided RdRp is widely present among fungal viruses, despite the belief that all RNA viruses encode RdRp as a single gene. These findings illustrate the previously hidden diversity and evolution of RNA viruses, and prompt reconsideration of the structural plasticity of RdRp. By highlighting the limitations of conventional surveillance methods for RNA viruses, we showcase the potential of FLDS technology to broaden current knowledge about these viruses.Author SummaryThe development of RNA-seq technology has facilitated the discovery of RNA viruses in all types of biological samples. However, it is technically difficult to detect highly novel viruses using RNA-seq. We successfully reconstructed the genomes of multiple novel fungal RNA viruses by screening host fungi using a new technology, FLDS. Surprisingly, we identified two viral species whose RNA-dependent RNA polymerase (RdRp) proteins were separately encoded on different genome segments, overturning the commonly accepted view of the positional unity of RdRp proteins in viral genomes. This new perspective on divided RdRp proteins should hasten the discovery of viruses with unique RdRp structures that have been overlooked, and further advance current knowledge and understanding of the diversity and evolution of RNA viruses.

scholarly journals Discovery of divided RdRp sequences and a hitherto unknown genomic complexity in fungal viruses

2020 ◽  
Author(s):  
Yuto Chiba ◽  
Sayoko Oiki ◽  
Takashi Yaguchi ◽  
Syun-Ichi Urayama ◽  
Daisuke Hagiwara
Keyword(s):  
Rna Viruses ◽  
Rna Virus ◽  
Single Gene ◽  
Virus Infections ◽  
Independent Manner ◽  
Viral Genomes ◽  
Genomic Complexity ◽  
True Diversity ◽  
Genomic Regions ◽  
Fungal Viruses

Abstract By identifying variations in viral RNA genomes, cutting-edge metagenome technology has potential to reshape current concepts about the evolution of RNA viruses. This technology, however, cannot process low-homology genomic regions properly, leaving the true diversity of RNA viruses unappreciated. To overcome this technological limitation, we applied an advanced method, Fragmented and Primer-Ligated Double-stranded (ds) RNA Sequencing (FLDS), to screen RNA viruses from 155 fungal isolates, which allowed us to obtain complete viral genomes in a homology-independent manner. We created a high-quality catalog of 19 RNA viruses (12 viral species) that infect Aspergillus isolates. Among them, nine viruses were not detectable by the conventional methodology involving agarose gel electrophoresis of dsRNA, a hallmark of RNA virus infections. Segmented genome structures were determined in 42% of the viruses. Some RNA viruses had novel genome architectures; one contained a dual methyltransferase domain and another had a separated RNA-dependent RNA polymerase (RdRp) gene. A virus from a different fungal taxon (Pyricularia) had an RdRp sequence that was separated on different segments, suggesting that a divided RdRp is widely present among fungal viruses, despite the belief that all RNA viruses encode RdRp as a single gene. These findings illustrate the previously hidden diversity and evolution of RNA viruses, and prompt reconsideration of the structural plasticity of RdRp.

scholarly journals RNA-Dependent RNA Polymerase from Heterobasidion RNA Virus 6 Is an Active Replicase In Vitro

Viruses ◽  
2021 ◽  
Vol 13 (9) ◽  
pp. 1738
Author(s):  
Alesia A. Levanova ◽  
Eeva J. Vainio ◽  
Jarkko Hantula ◽  
Minna M. Poranen
Keyword(s):  
Rna Polymerase ◽  
Rna Synthesis ◽  
Rna Virus ◽  
Polymerization Reaction ◽  
Rna Dependent Rna Polymerase ◽  
Independent Manner ◽  
Nucleotidyl Transferase ◽  
Rna Polymerization ◽  
The Family

Heterobasidion RNA virus 6 (HetRV6) is a double-stranded (ds)RNA mycovirus and a member of the recently established genus Orthocurvulavirus within the family Orthocurvulaviridae. The purpose of the study was to determine the biochemical requirements for RNA synthesis catalyzed by HetRV6 RNA-dependent RNA polymerase (RdRp). HetRV6 RdRp was expressed in Escherichia coli and isolated to near homogeneity using liquid chromatography. The enzyme activities were studied in vitro using radiolabeled UTP. The HetRV6 RdRp was able to initiate RNA synthesis in a primer-independent manner using both virus-related and heterologous single-stranded (ss)RNA templates, with a polymerization rate of about 46 nt/min under optimal NTP concentration and temperature. NTPs with 2′-fluoro modifications were also accepted as substrates in the HetRV6 RdRp-catalyzed RNA polymerization reaction. HetRV6 RdRp transcribed viral RNA genome via semi-conservative mechanism. Furthermore, the enzyme demonstrated terminal nucleotidyl transferase (TNTase) activity. Presence of Mn2+ was required for the HetRV6 RdRp catalyzed enzymatic activities. In summary, our study shows that HetRV6 RdRp is an active replicase in vitro that can be potentially used in biotechnological applications, molecular biology, and biomedicine.

scholarly journals Recombination Located over 2A-2B Junction Ribosome Frameshifting Region of Saffold Cardiovirus

Viruses ◽  
2018 ◽  
Vol 10 (10) ◽  
pp. 520 ◽  
Author(s):  
Antônio da Costa ◽  
Adriana Luchs ◽  
Flávio Milagres ◽  
Shirley Komninakis ◽  
Danielle Gill ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Acute Gastroenteritis ◽  
Empirical Studies ◽  
Full Length ◽  
Evolutionary Analysis ◽  
Ribosomal Frameshift ◽  
Rna Dependent Rna Polymerase ◽  
Recombination Hotspot ◽  
Viral Genomes ◽  
Full Length Genome

Here we report the nearly full-length genome of a recombinant Saffold virus strain (SAFV-BR-193) isolated from a child with acute gastroenteritis. Evolutionary analysis performed using all available near-full length Saffold picornavirus genomes showed that the breakpoint found in the Brazilian strain (SAFV-BR-193) is indeed a recombination hotspot. Notably, this hotspot is located just one nucleotide after the ribosomal frameshift GGUUUUU motif in the SAFV genome. Empirical studies will be necessary to determine if this motif also affects the binding affinity of RNA-dependent RNA-polymerase (RdRp) and therefore increases the changes of RdRp swap between molecules during the synthesis of viral genomes.

scholarly journals RNA Recombination In Vivo in the Absence of Viral Replication

2004 ◽  
Vol 78 (12) ◽  
pp. 6271-6281 ◽  
Author(s):  
Andreas Gallei ◽  
Alexander Pankraz ◽  
Heinz-Jürgen Thiel ◽  
Paul Becher
Keyword(s):  
Rna Viruses ◽  
Rna Recombination ◽  
Rna Dependent Rna Polymerase ◽  
Viral Genomes ◽  
Nonhomologous Recombination ◽  
Viral Rdrp ◽  
Recombination System ◽  
Independent Mechanism ◽  
Overlapping Transcripts

ABSTRACT To study fundamental aspects of RNA recombination, an in vivo RNA recombination system was established. This system allowed the efficient generation of recombinant cytopathogenic pestiviruses after transfection of synthetic, nonreplicatable, subgenomic transcripts in cells infected with a replicating noncytopathogenic virus. Studies addressing the interplay between RNA recombination and replication revealed that cotransfection of noninfected cells with various pairs of nonreplicatable RNA derivatives also led to the emergence of recombinant viral genomes. Remarkably, homologous and nonhomologous recombination occurred between two overlapping transcripts, each lacking different essential parts of the viral RNA-dependent RNA polymerase (RdRp) gene. Apart from the generally accepted viral replicative copy choice recombination, our results prove the existence of a viral RdRp-independent mechanism of RNA recombination that occurs in vivo. It appears likely that such a mechanism not only contributes to the evolution of RNA viruses but also leads to the generation of recombinant cellular RNAs.

scholarly journals Late Male-Killing Viruses in Homona magnanima Identified as Osugoroshi Viruses, Novel Members of Partitiviridae

2021 ◽  
Vol 11 ◽  
Author(s):  
Ryosuke Fujita ◽  
Maki N. Inoue ◽  
Takumi Takamatsu ◽  
Hiroshi Arai ◽  
Mayu Nishino ◽  
...  
Keyword(s):  
Mathematical Modeling ◽  
Rna Polymerase ◽  
Rna Viruses ◽  
Rna Virus ◽  
Horizontal Transmission ◽  
Rna Dependent Rna Polymerase ◽  
Double Stranded Rna ◽  
First Report ◽  
Male Killing ◽  
Male Specific

Late male-killing, a male-specific death after hatching, is a unique phenotype found in Homona magnanima, oriental tea tortrix. The male-killing agent was suspected to be an RNA virus, but details were unknown. We herein successfully isolated and identified the putative male-killing virus as Osugoroshi viruses (OGVs). The three RNA-dependent RNA polymerase genes detected were phylogenetically related to Partitiviridae, a group of segmented double-stranded RNA viruses. Purified dsRNA from a late male-killing strain of H. magnanima revealed 24 segments, in addition to the RdRps, with consensus terminal sequences. These segments included the previously found male-killing agents MK1068 (herein OGV-related RNA16) and MK1241 (OGV-related RNA7) RNAs. Ultramicroscopic observation of purified virions, which induced late male-killing in the progeny of injected moths, showed sizes typical of Partitiviridae. Mathematical modeling showed the importance of late male-killing in facilitating horizontal transmission of OGVs in an H. magnanima population. This study is the first report on the isolation of partiti-like virus from insects, and one thought to be associated with late male-killing, although the viral genomic contents and combinations in each virus are still unknown.

Targeting Viral RNA-Dependent RNA Polymerases as an Antiviral Therapy

2021 ◽  
Vol 28 ◽  
Author(s):  
Daniel Miranda ◽  
David Jesse Sanchez
Keyword(s):  
Clinical Trials ◽  
Global Health ◽  
Rna Polymerase ◽  
Biomedical Research ◽  
Rna Viruses ◽  
Standard Of Care ◽  
Rna Dependent Rna Polymerase ◽  
Viral Pathogens ◽  
Relevant Topic ◽  
Target Rna

Abstract: Progressive globalization of our society brings not only worldwide integration, it increases and promotes our exposure to new viral pathogens with evident impacts on our global health. Especially with the emergence of SARS-CoV-2, our biomedical research infrastructure has never been more compelled to rapidly develop antiviral regimens that demonstrate improved efficacy against these pathogens. Here we showcase 3 poignant antivirals against the lucrative target, RNA-dependent RNA polymerase (RdRP) of RNA viruses – a timely and relevant topic given the present efforts against COVID-19. While effective drug designs against RdRP are important, their benefit and potential as a standard of care truly relies on them standing out in well-designed clinical trials.

scholarly journals Products of Oxidative Guanine Damage Form Base Pairs with Guanine

2020 ◽  
Vol 21 (20) ◽  
pp. 7645
Author(s):  
Katsuhito Kino ◽  
Taishu Kawada ◽  
Masayo Hirao-Suzuki ◽  
Masayuki Morikawa ◽  
Hiroshi Miyazawa
Keyword(s):  
Carboxylic Acid ◽  
Rna Polymerase ◽  
Rna Viruses ◽  
Antiviral Agents ◽  
Present Knowledge ◽  
Base Pairs ◽  
Rna Dependent Rna Polymerase ◽  
Novel Coronavirus

Among the natural bases, guanine is the most oxidizable base. The damage caused by oxidation of guanine, commonly referred to as oxidative guanine damage, results in the formation of several products, including 2,5-diamino-4H-imidazol-4-one (Iz), 2,2,4-triamino-5(2H)-oxazolone (Oz), guanidinoformimine (Gf), guanidinohydantoin/iminoallantoin (Gh/Ia), spiroiminodihydantoin (Sp), 5-carboxamido-5-formamido-2-iminohydantoin (2Ih), urea (Ua), 5-guanidino-4-nitroimidazole (NI), spirodi(iminohydantoin) (5-Si and 8-Si), triazine, the M+7 product, other products by peroxynitrite, alkylated guanines, and 8,5′-cyclo-2′-deoxyguanosine (cG). Herein, we summarize the present knowledge about base pairs containing the products of oxidative guanine damage and guanine. Of these products, Iz is involved in G-C transversions. Oz, Gh/Ia, and Sp form preferably Oz:G, Gh/Ia:G, and Sp:G base pairs in some cases. An involvement of Gf, 2Ih, Ua, 5-Si, 8-Si, triazine, the M+7 product, and 4-hydroxy-2,5-dioxo-imidazolidine-4-carboxylic acid (HICA) in G-C transversions requires further experiments. In addition, we describe base pairs that target the RNA-dependent RNA polymerase (RdRp) of RNA viruses and describe implications for the 2019 novel coronavirus (SARS-CoV-2): When products of oxidative guanine damage are adapted for the ribonucleoside analogs, mimics of oxidative guanine damages, which can form base pairs, may become antiviral agents for SARS-CoV-2.

scholarly journals Viral discovery and diversity in trypanosomatid protozoa with a focus on relatives of the human parasiteLeishmania

2017 ◽  
Vol 115 (3) ◽  
pp. E506-E515 ◽  
Author(s):  
Danyil Grybchuk ◽  
Natalia S. Akopyants ◽  
Alexei Y. Kostygov ◽  
Aleksandras Konovalovas ◽  
Lon-Fye Lye ◽  
...  
Keyword(s):  
Visceral Leishmaniasis ◽  
Rna Polymerase ◽  
Human Disease ◽  
Rna Viruses ◽  
Biological Role ◽  
Viral Diversity ◽  
Rna Dependent Rna Polymerase ◽  
Human Parasite ◽  
Viral Lineage

Knowledge of viral diversity is expanding greatly, but many lineages remain underexplored. We surveyed RNA viruses in 52 cultured monoxenous relatives of the human parasiteLeishmania(CrithidiaandLeptomonas), as well as plant-infectingPhytomonas.Leptomonas pyrrhocoriswas a hotbed for viral discovery, carrying a virus (Leptomonas pyrrhocoris ostravirus 1) with a highly divergent RNA-dependent RNA polymerase missed by conventional BLAST searches, an emergent clade of tombus-like viruses, and an example of viral endogenization. A deep-branching clade of trypanosomatid narnaviruses was found, notable asLeptomonas seymouribearing Narna-like virus 1 (LepseyNLV1) have been reported in cultures recovered from patients with visceral leishmaniasis. A deep-branching trypanosomatid viral lineage showing strong affinities to bunyaviruses was termed “Leishbunyavirus” (LBV) and judged sufficiently distinct to warrant assignment within a proposed family termed “Leishbunyaviridae.” Numerous relatives of trypanosomatid viruses were found in insect metatranscriptomic surveys, which likely arise from trypanosomatid microbiota. Despite extensive sampling we found no relatives of the totivirusLeishmaniavirus(LRV1/2), implying that it was acquired at about the same time theLeishmaniabecame able to parasitize vertebrates. As viruses were found in over a quarter of isolates tested, many more are likely to be found in the >600 unsurveyed trypanosomatid species. Viral loss was occasionally observed in culture, providing potentially isogenic virus-free lines enabling studies probing the biological role of trypanosomatid viruses. These data shed important insights on the emergence of viruses within an important trypanosomatid clade relevant to human disease.

scholarly journals Protein Nucleotidylylation in +ssRNA Viruses

Viruses ◽  
2021 ◽  
Vol 13 (8) ◽  
pp. 1549
Author(s):  
Alice-Roza Eruera ◽  
Alice M. McSweeney ◽  
Geena M. McKenzie-Goldsmith ◽  
Vernon K. Ward
Keyword(s):  
Life Cycle ◽  
Rna Polymerase ◽  
Viral Replication ◽  
Viral Protein ◽  
Rna Viruses ◽  
Replication Initiation ◽  
Genome Replication ◽  
Rna Dependent Rna Polymerase ◽  
Nucleotidyl Transferase ◽  
Initiation Of Translation

Nucleotidylylation is a post-transcriptional modification important for replication in the picornavirus supergroup of RNA viruses, including members of the Caliciviridae, Coronaviridae, Picornaviridae and Potyviridae virus families. This modification occurs when the RNA-dependent RNA polymerase (RdRp) attaches one or more nucleotides to a target protein through a nucleotidyl-transferase reaction. The most characterized nucleotidylylation target is VPg (viral protein genome-linked), a protein linked to the 5′ end of the genome in Caliciviridae, Picornaviridae and Potyviridae. The nucleotidylylation of VPg by RdRp is a critical step for the VPg protein to act as a primer for genome replication and, in Caliciviridae and Potyviridae, for the initiation of translation. In contrast, Coronaviridae do not express a VPg protein, but the nucleotidylylation of proteins involved in replication initiation is critical for genome replication. Furthermore, the RdRp proteins of the viruses that perform nucleotidylylation are themselves nucleotidylylated, and in the case of coronavirus, this has been shown to be essential for viral replication. This review focuses on nucleotidylylation within the picornavirus supergroup of viruses, including the proteins that are modified, what is known about the nucleotidylylation process and the roles that these modifications have in the viral life cycle.

scholarly journals NeoRdRp: A comprehensive dataset for identifying RNA-dependent RNA polymerase of various RNA viruses from metatranscriptomic data

2022 ◽  
Author(s):  
Shoichi Sakaguchi ◽  
Syun-ichi Urayama ◽  
Yoshihiro Takaki ◽  
Hong Wu ◽  
Youichi Suzuki ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Viruses ◽  
Rna Virus ◽  
Sequence Similarity ◽  
Virus Detection ◽  
Detection Methods ◽  
Amino Acid Sequence Similarity ◽  
Sequencing Data ◽  
Rna Dependent Rna Polymerase ◽  
Multiple Sequence

RNA viruses are distributed in various environments, and most RNA viruses have been recently identified by metatranscriptome sequencing. However, due to the high nucleotide diversity of RNA viruses, it is still challenging to identify their sequences. Therefore, this study generated a dataset of RNA-dependent RNA polymerase (RdRp) domains essential for all RNA viruses belonging to Orthornavirae. Also, the collected genes with RdRp domains from various RNA viruses were clustered by amino acid sequence similarity. For each cluster, a multiple sequence alignment was generated, and a hidden Markov model (HMM) profile was created if the number of sequences was greater than five. Using the 1,467 HMM profiles, we detected RdRp domains in the RefSeq RNA virus sequences, combined the hit sequences with the RdRp domains, and reconstructed the HMM profiles. As a result, 2,234 HMM profiles were generated from 12,316 RdRp domain sequences, and the dataset was named NeoRdRp. Additionally, using the UniProt dataset, we confirmed that almost all NeoRdRp HMM profiles could specifically detect RdRps in Orthornavirae. Furthermore, we compared the NeoRdRp dataset with two previously reported RNA virus detection methods to detect RNA virus sequences from metatranscriptome sequencing data. Our methods can identify most of the RNA viruses in the datasets; however, some RNA viruses were not detected, similar to the other two methods. The NeoRdRp can be improved by repeatedly adding new RdRp sequences and can be expected to be widely applied as a system for detecting various RNA viruses from metatranscriptome data.

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