scholarly journals A Newly Identified Virus in the Family Potyviridae Encodes Two Leader Cysteine Proteases in Tandem That Evolved Contrasting RNA Silencing Suppression Functions

2020 ◽  
Vol 95 (1) ◽  
Author(s):  
Li Qin ◽  
Wentao Shen ◽  
Zhongfa Tang ◽  
Weiyao Hu ◽  
Lingna Shangguan ◽  
...  
Keyword(s):  
Rna Silencing ◽  
Rna Viruses ◽  
Evolutionary Relationship ◽  
Cysteine Proteases ◽  
Terminal Region ◽  
Crop Damage ◽  
Virus Species ◽  
Protease Domain ◽  
The Family ◽  
Silencing Suppression

ABSTRACT Potyviridae is the largest family of plant-infecting RNA viruses and includes many agriculturally and economically important viral pathogens. The viruses in the family, known as potyvirids, possess single-stranded, positive-sense RNA genomes with polyprotein processing as a gene expression strategy. The N-terminal regions of potyvirid polyproteins vary greatly in sequence. Previously, we identified a novel virus species within the family, Areca palm necrotic spindle-spot virus (ANSSV), which was predicted to encode two cysteine proteases, HCPro1 and HCPro2, in tandem at the N-terminal region. Here, we present evidence showing self-cleavage activity of these two proteins and define their cis-cleavage sites. We demonstrate that HCPro2 is a viral suppressor of RNA silencing (VSR), and both the variable N-terminal and conserved C-terminal (protease domain) moieties have antisilencing activity. Intriguingly, the N-terminal region of HCPro1 also has RNA silencing suppression activity, which is, however, suppressed by its C-terminal protease domain, leading to the functional divergence of HCPro1 and HCPro2 in RNA silencing suppression. Moreover, the deletion of HCPro1 or HCPro2 in a newly created infectious clone abolishes viral infection, and the deletion mutants cannot be rescued by addition of corresponding counterparts of a potyvirus. Altogether, these data suggest that the two closely related leader proteases of ANSSV have evolved differential and essential functions to concertedly maintain viral viability. IMPORTANCE The Potyviridae represent the largest group of known plant RNA viruses and account for more than half of the viral crop damage worldwide. The leader proteases of viruses within the family vary greatly in size and arrangement and play key roles during the infection. Here, we experimentally demonstrate the presence of a distinct pattern of leader proteases, HCPro1 and HCPro2 in tandem, in a newly identified member within the family. Moreover, HCPro1 and HCPro2, which are closely related and typically characterized with a short size, have evolved contrasting RNA silencing suppression activity and seem to function in a coordinated manner to maintain viral infectivity. Altogether, the new knowledge fills a missing piece in the evolutionary relationship history of potyvirids and improves our understanding of the diversification of potyvirid genomes.

scholarly journals RNA Silencing Suppression by a Second Copy of the P1 Serine Protease of Cucumber Vein Yellowing Ipomovirus, a Member of the Family Potyviridae That Lacks the Cysteine Protease HCPro

2006 ◽  
Vol 80 (20) ◽  
pp. 10055-10063 ◽  
Author(s):  
Adrian Valli ◽  
Ana Montserrat Martín-Hernández ◽  
Juan José López-Moya ◽  
Juan Antonio García
Keyword(s):  
Rna Silencing ◽  
Fluorescent Protein ◽  
Serine Proteinase ◽  
Plum Pox Virus ◽  
Coding Region ◽  
Long Distance ◽  
Systemic Spread ◽  
The Family ◽  
Silencing Suppression ◽  
Green Fluorescent

ABSTRACT The P1 protein of viruses of the family Potyviridae is a serine proteinase, which is highly variable in length and sequence, and its role in the virus infection cycle is not clear. One of the proposed activities of P1 is to assist HCPro, the product that viruses of the genus Potyvirus use to counteract antiviral defense mediated by RNA silencing. Indeed, an HCPro-coding region is present in all the genomes of members of the genera Potyvirus, Rymovirus, and Tritimovirus that have been sequenced. However, it was recently reported that a sequence coding for HCPro is lacking in the genome of Cucumber vein yellowing virus (CVYV), a member of the genus Ipomovirus, the fourth monopartite genus of the family. In this study, we provide further evidence that P1 enhances the activity of HCPro in members of the genus Potyvirus and show that it is duplicated in the ipomovirus CVYV. The two CVYV P1 copies are arranged in tandem, and the second copy (P1b) has RNA silencing suppression activity. CVYV P1b suppressed RNA silencing induced either by sense green fluorescent protein (GFP) mRNA or by a GFP inverted repeat RNA, indicating that CVYV P1b acts downstream of the formation of double-stranded RNA. CVYV P1b also suppressed local silencing in agroinfiltrated patches of transgenic Nicotiana benthamiana line 16c and delayed its propagation to the neighboring cells. However, neither the short-distance nor long-distance systemic spread of silencing of the GFP transgene was completely blocked by CVYV P1b. CVYV P1b and P1-HCPro from the potyvirus Plum pox virus showed very similar behaviors in all the assays carried out, suggesting that evolution has found a way to counteract RNA silencing by similar mechanisms using very different proteins in viruses of the same family.

scholarly journals Strawberry Mottle Virus (Family Secoviridae, Order Picornavirales) Encodes a Novel Glutamic Protease To Process the RNA2 Polyprotein at Two Cleavage Sites

2018 ◽  
Vol 93 (5) ◽  
Author(s):  
Krin S. Mann ◽  
Joan Chisholm ◽  
Hélène Sanfaçon
Keyword(s):  
Cysteine Proteases ◽  
Terminal Region ◽  
Mottle Virus ◽  
Cleavage Sites ◽  
Virus Family ◽  
Positive Strand ◽  
Viral Proteases ◽  
Bipartite Genome ◽  
The Family ◽  
Positive Strand Rna

ABSTRACT Strawberry mottle virus (SMoV) belongs to the family Secoviridae (order Picornavirales) and has a bipartite genome with each RNA encoding one polyprotein. All characterized secovirids encode a single protease related to the picornavirus 3C protease. The SMoV 3C-like protease was previously shown to cut the RNA2 polyprotein (P2) at a single site between the predicted movement protein and coat protein (CP) domains. However, the SMoV P2 polyprotein includes an extended C-terminal region with a coding capacity of up to 70 kDa downstream of the presumed CP domain, an unusual characteristic for this family. In this study, we identified a novel cleavage event at a P↓AFP sequence immediately downstream of the CP domain. Following deletion of the PAFP sequence, the polyprotein was processed at or near a related PKFP sequence 40 kDa further downstream, defining two protein domains in the C-terminal region of the P2 polyprotein. Both processing events were dependent on a novel protease domain located between the two cleavage sites. Mutagenesis of amino acids that are conserved among isolates of SMoV and of the related Black raspberry necrosis virus did not identify essential cysteine, serine, or histidine residues, suggesting that the RNA2-encoded SMoV protease is not related to serine or cysteine proteases of other picorna-like viruses. Rather, two highly conserved glutamic acid residues spaced by 82 residues were found to be strictly required for protease activity. We conclude that the processing of SMoV polyproteins requires two viral proteases, the RNA1-encoded 3C-like protease and a novel glutamic protease encoded by RNA2. IMPORTANCE Many viruses encode proteases to release mature proteins and intermediate polyproteins from viral polyproteins. Polyprotein processing allows regulation of the accumulation and activity of viral proteins. Many viral proteases also cleave host factors to facilitate virus infection. Thus, viral proteases are key virulence factors. To date, viruses with a positive-strand RNA genome are only known to encode cysteine or serine proteases, most of which are related to the cellular papain, trypsin, or chymotrypsin proteases. Here, we characterize the first glutamic protease encoded by a plant virus or by a positive-strand RNA virus. The novel glutamic protease is unique to a few members of the family Secoviridae, suggesting that it is a recent acquisition in the evolution of this family. The protease does not resemble known cellular proteases. Rather, it is predicted to share structural similarities with a family of fungal and bacterial glutamic proteases that adopt a lectin fold.

scholarly journals Functional characterization and subcellular localization of the 16K cysteine-rich suppressor of gene silencing protein of tobacco rattle virus

2008 ◽  
Vol 89 (7) ◽  
pp. 1748-1758 ◽  
Author(s):  
Walid Ghazala ◽  
Angelika Waltermann ◽  
Ruth Pilot ◽  
Stephan Winter ◽  
Mark Varrelmann
Keyword(s):  
Nuclear Localization ◽  
Rna Silencing ◽  
Fluorescent Protein ◽  
Functional Characterization ◽  
Tobacco Rattle Virus ◽  
Terminal Region ◽  
Laser Scanning Microscopy ◽  
Nuclear Targeting ◽  
Nuclear Localization Signals ◽  
Silencing Suppression

The 16 kDa cysteine-rich protein (16K) of tobacco rattle virus (TRV) is known to partially suppress RNA silencing in Drosophila cells. In this study, we show that 16K suppresses RNA silencing in green fluorescent protein (GFP)-transgenic Nicotiana benthamiana plants using an Agrobacterium-mediated transient assay. 16K slightly reduced the accumulation of short interfering RNAs (siRNA) of GFP, suggesting that the protein may interfere with the initiation and/or maintenance of RNA silencing. Deletion of either the N- or C-terminal part of 16K indicated that the entire 16K open reading frame (ORF) is necessary for its silencing suppression function. Pentapeptide insertion scanning mutagenesis (PSM) revealed that only two short regions of 16K tolerated five extra amino acid insertions without considerable reduction in its silencing suppression function. The tolerant regions coincide with sequence variability between tobravirus cysteine-rich proteins, indicating a strong functional and/or structural conservation of TRV 16K. Confocal laser scanning microscopy of transiently expressed 16K fusions to red fluorescent protein (RFP) revealed a predominant cytoplasmic localization and, in addition, a nuclear localization. In contrast, fusions of RFP with the N-terminal region of 16K localized exclusively to the cytoplasm, whereas fusions between RFP and the C-terminal region of 16K displayed an exclusive nuclear localization. Further analysis of 16K-derived peptide fusions demonstrated that the 16K C-terminal region contained at least two functional bipartite nuclear localization signals which were independently capable of nuclear targeting.

scholarly journals Variability in the level of RNA silencing suppression caused by triple gene block protein 1 (TGBp1) from various potexviruses during infection

2009 ◽  
Vol 90 (4) ◽  
pp. 1014-1024 ◽  
Author(s):  
Hiroko Senshu ◽  
Johji Ozeki ◽  
Ken Komatsu ◽  
Masayoshi Hashimoto ◽  
Kouji Hatada ◽  
...  
Keyword(s):  
Virus Infection ◽  
Rna Silencing ◽  
Triple Gene Block ◽  
Plant Viruses ◽  
Virus Species ◽  
Suppressor Activity ◽  
Gene Block ◽  
Gfp Fluorescence ◽  
Silencing Suppression ◽  
Triple Gene Block Protein

RNA silencing is an important defence mechanism against virus infection, and many plant viruses encode RNA silencing suppressors as a counter defence. In this study, we analysed the RNA silencing suppression ability of multiple virus species of the genus Potexvirus. Nicotiana benthamiana plants exhibiting RNA silencing of a green fluorescent protein (GFP) transgene showed reversal of GFP fluorescence when systemically infected with potexviruses. However, the degree of GFP fluorescence varied among potexviruses. Agrobacterium-mediated transient expression assay in N. benthamiana leaves demonstrated that the triple gene block protein 1 (TGBp1) encoded by these potexviruses has drastically different levels of silencing suppressor activity, and these differences were directly related to variations in the silencing suppression ability during virus infection. These results suggest that suppressor activities differ even among homologous proteins encoded by viruses of the same genus, and that TGBp1 contributes to the variation in the level of RNA silencing suppression by potexviruses. Moreover, we investigated the effect of TGBp1 encoded by Plantago asiatica mosaic virus (PlAMV), which exhibited a strong suppressor activity, on the accumulation of microRNA, virus genomic RNA and virus-derived small interfering RNAs.

scholarly journals Analysis of gene content in sweet potato chlorotic stunt virus RNA1 reveals the presence of the p22 RNA silencing suppressor in only a few isolates: implications for viral evolution and synergism

2008 ◽  
Vol 89 (2) ◽  
pp. 573-582 ◽  
Author(s):  
Wilmer J. Cuellar ◽  
Fred Tairo ◽  
Jan F. Kreuze ◽  
Jari P. T. Valkonen
Keyword(s):  
Sweet Potato ◽  
Rna Silencing ◽  
Gene Content ◽  
Gene Gain ◽  
Virus Species ◽  
Silencing Suppressor ◽  
Virus Family ◽  
Rna Silencing Suppressor ◽  
African Strain ◽  
The Family

Sweet potato chlorotic stunt virus (genus Crinivirus) belongs to the family Closteroviridae, members of which have a conserved overall genomic organization but are variable in gene content. In the bipartite criniviruses, heterogeneity is pronounced in the 3′-proximal region of RNA1, which in sweet potato chlorotic stuat virus (SPCSV) encodes two novel proteins, RNase3 (RNase III endonuclease) and p22 (RNA silencing suppressor). This study showed that two Ugandan SPCSV isolates contained the p22 gene, in contrast to three isolates of the East African strain from Tanzania and Peru and an isolate of the West African strain from Israel, which were missing a 767 nt fragment of RNA1 that included the p22 gene. Regardless of the presence of p22, all tested SPCSV isolates acted synergistically with potyvirus sweet potato feathery mottle virus (SPFMV; genus Potyvirus, family Potyviridae) in co-infected sweetpotato plants (Ipomoea batatas), which greatly enhanced SPFMV titres and caused severe sweetpotato virus disease (SPVD). Therefore, the results indicate that any efforts to engineer pathogen-derived RNA silencing-based resistance to SPCSV and SPVD in sweetpotato should not rely on p22 as the transgene. The data from this study demonstrate that isolates of this virus species can vary in the genes encoding RNA silencing suppressor proteins. This study also provides the first example of intraspecific variability in gene content of the family Closteroviridae and may be a new example of the recombination-mediated gene gain that is characteristic of virus evolution in this virus family.

scholarly journals Heterologous RNA-silencing suppressors from both plant- and animal-infecting viruses support plum pox virus infection

2012 ◽  
Vol 93 (7) ◽  
pp. 1601-1611 ◽  
Author(s):  
Varvara I. Maliogka ◽  
María Calvo ◽  
Alberto Carbonell ◽  
Juan Antonio García ◽  
Adrian Valli
Keyword(s):  
Viral Infection ◽  
Rna Silencing ◽  
Influenza A ◽  
Plum Pox Virus ◽  
Ns1 Protein ◽  
Multifunctional Protein ◽  
Highly Sensitive ◽  
The Family ◽  
Silencing Suppression ◽  
Insight Into

HCPro, the RNA-silencing suppressor (RSS) of viruses belonging to the genus Potyvirus in the family Potyviridae, is a multifunctional protein presumably involved in all essential steps of the viral infection cycle. Recent studies have shown that plum pox potyvirus (PPV) HCPro can be replaced successfully by cucumber vein yellowing ipomovirus P1b, a sequence-unrelated RSS from a virus of the same family. In order to gain insight into the requirement of a particular RSS to establish a successful potyviral infection, we tested the ability of different heterologous RSSs from both plant- and animal-infecting viruses to substitute for HCPro. Making use of engineered PPV chimeras, we show that PPV HCPro can be replaced functionally by some, but not all, unrelated RSSs, including the NS1 protein of the mammal-infecting influenza A virus. Interestingly, the capacity of a particular RSS to replace HCPro does not correlate strictly with its RNA silencing-suppression strength. Altogether, our results suggest that not all suppression strategies are equally suitable for efficient escape of PPV from the RNA-silencing machinery. The approach followed here, based on using PPV chimeras in which an under-consideration RSS substitutes for HCPro, could further help to study the function of diverse RSSs in a ‘highly sensitive’ RNA-silencing context, such as that taking place in plant cells during the process of a viral infection.

scholarly journals Wheat streak mosaic virus P1 Binds to dsRNAs without Size and Sequence Specificity and a GW Motif Is Crucial for Suppression of RNA Silencing

Viruses ◽  
2019 ◽  
Vol 11 (5) ◽  
pp. 472 ◽  
Author(s):  
Adarsh K. Gupta ◽  
Satyanarayana Tatineni
Keyword(s):  
Mosaic Virus ◽  
Rna Silencing ◽  
Fluorescent Protein ◽  
Wheat Streak Mosaic Virus ◽  
Terminal Region ◽  
Single Amino Acid ◽  
Sequence Specificity ◽  
Independent Manner ◽  
Silencing Suppression

Wheat streak mosaic virus (WSMV; genus Tritimovirus; family Potyviridae) is an economically important virus infecting wheat in the Great Plains region of the USA. Previously, we reported that the P1 protein of WSMV acts as a viral suppressor of RNA silencing. In this study, we delineated the minimal region of WSMV P1 and examined its mechanisms in suppression of RNA silencing. We found that the 25 N-terminal amino acids are dispensable, while deletion of a single amino acid at the C-terminal region completely abolished the RNA silencing suppression activity of P1. Electrophoretic mobility shift assays with in vitro expressed P1 revealed that the P1 protein formed complexes with green fluorescent protein-derived 180-nt dsRNA and 21 and 24-nt ds-siRNAs, and WSMV coat protein-specific 600-nt dsRNA. These data suggest that the P1 protein of WSMV binds to dsRNAs in a size- and sequence-independent manner. Additionally, in vitro dicing assay with human Dicer revealed that the P1 protein efficiently protects dsRNAs from processing by Dicer into siRNAs, by forming complexes with dsRNA. Sequence comparison of P1-like proteins from select potyvirid species revealed that WSMV P1 harbors a glycine-tryptophan (GW) motif at the C-terminal region. Disruption of GW motif in WSMV P1 through W303A mutation resulted in loss of silencing suppression function and pathogenicity enhancement, and abolished WSMV viability. These data suggest that the mechanisms of suppression of RNA silencing of P1 proteins of potyvirid species appear to be broadly conserved in the family Potyviridae.

scholarly journals The role of chromosomal fusion in the karyotypic evolution of the genus Ageneiosus (Siluriformes: Auchenipteridae)

2013 ◽  
Vol 11 (2) ◽  
pp. 327-334 ◽  
Author(s):  
Roberto Laridondo Lui ◽  
Daniel Rodrigues Blanco ◽  
Juliana de Fatima Martinez ◽  
Vladimir Pavan Margarido ◽  
Paulo Cesar Venere ◽  
...  
Keyword(s):  
Chromosome Pair ◽  
5S Rdna ◽  
Proximal Region ◽  
Terminal Region ◽  
Metacentric Chromosome ◽  
South American ◽  
Submetacentric Chromosome ◽  
Araguaia River ◽  
The Family ◽  
Preferential Location

Ageneiosus is the most widely distributed genus of the family Auchenipteridae among South American river basins. Although chromosome studies in the family are scarce, this genus has the largest number of analyzed species, with 2n = 54 to 56 chromosomes, differing from the rest of the family (2n = 58). This study aimed to analyze Ageneiosus inermis from the Araguaia River basin. The diploid number found was of 56 chromosomes. Heterochromatin was allocated in terminal region of most chromosomes, plus a pericentromeric heterochromatic block in pair 1, a pair distinguished by size in relation to other chromosomes pairs. AgNORs were detected in only one submetacentric chromosome pair, which was confirmed by FISH. 5S rDNA was present in only one metacentric chromosome pair. Hybridization with [TTAGGG]n sequence marked the telomeres of all chromosomes, in addition to an ITS in the proximal region of the short arm of pair 1. The repetitive [GATA]n sequence was dispersed, with preferential location in terminal region of the chromosomes. Ageneiosus has a genomic organization somewhat different when compared to other Auchenipteridae species. Evidences indicate that a chromosomal fusion originated the first metacentric chromosome pair in A. inermis, rearrangement which may be a basal event for the genus

Flaviviruses excluding dengue

2020 ◽  
pp. 830-845
Author(s):  
Shannan Lee Rossi ◽  
Nikos Vasilakis
Keyword(s):  
Japanese Encephalitis Virus ◽  
Host Range ◽  
Yellow Fever ◽  
Japanese Encephalitis ◽  
Human Infection ◽  
Virus Species ◽  
Human Pathogens ◽  
West Nile ◽  
The Family ◽  
Large Host

The family Flaviviridae currently consists of four recognized genera: Flavivirus, Pestivirus, Hepacivirus, and Pegivirus. Although members of the family have a large host range that includes both vertebrates and invertebrates, only members of the genus Flavivirus are known as arboviruses, vectored either by mosquitoes or ticks. The remaining genera in the family are exclusively found in mammals, and their diversity has greatly expanded with recent virus discoveries. The genus Flavivirus comprises 92 virus species, of which over 40 can cause human infection. Many of these include important human pathogens such as Zika, dengue, yellow fever, West Nile, and Japanese encephalitis virus.

scholarly journals Tracing the Lineage of Two Traits Associated with the Coat Protein of the Tombusviridae: Silencing Suppression and HR Elicitation in Nicotiana Species

Viruses ◽  
2019 ◽  
Vol 11 (7) ◽  
pp. 588 ◽  
Author(s):  
Mustafa Adhab ◽  
Carlos Angel ◽  
Andres Rodriguez ◽  
Mohammad Fereidouni ◽  
Lóránt Király ◽  
...  
Keyword(s):  
Phylogenetic Analysis ◽  
Structural Changes ◽  
Coat Proteins ◽  
Silencing Suppressor ◽  
Suppressor Activity ◽  
Nicotiana Species ◽  
Pathogen Effector ◽  
The Family ◽  
Silencing Suppression ◽  
Insight Into

In this paper we have characterized the lineage of two traits associated with the coat proteins (CPs) of the tombusvirids: Silencing suppression and HR elicitation in Nicotiana species. We considered that the tombusvirid CPs might collectively be considered an effector, with the CP of each CP-encoding species comprising a structural variant within the family. Thus, a phylogenetic analysis of the CP could provide insight into the evolution of a pathogen effector. The phylogeny of the CP of tombusvirids indicated that CP representatives of the family could be divided into four clades. In two separate clades the CP triggered a hypersensitive response (HR) in Nicotiana species of section Alatae but did not have silencing suppressor activity. In a third clade the CP had a silencing suppressor activity but did not have the capacity to trigger HR in Nicotiana species. In the fourth clade, the CP did not carry either function. Our analysis illustrates how structural changes that likely occurred in the CP effector of progenitors of the current genera led to either silencing suppressor activity, HR elicitation in select Nicotiana species, or neither trait.

Sign in / Sign up

Export Citation Format

Share Document