Peste des petits ruminants(PPR) is a highly contagious transboundary animal disease with a severe socio-economic impact on the livestock industry, particularly in poor countries where it is endemic. Full understanding...Peste des petits ruminants(PPR) is a highly contagious transboundary animal disease with a severe socio-economic impact on the livestock industry, particularly in poor countries where it is endemic. Full understanding of PPR virus(PPRV)pathobiology and molecular biology is critical for effective control and eradication of the disease. To achieve these goals,establishment of stable reverse genetics systems for PPRV would play a key role. Unfortunately, this powerful technology remains less accessible and poorly documented for PPRV. In this review, we discussed the current status of PPRV reverse genetics as well as the recent innovations and advances in the reverse genetics of other non-segmented negative-sense RNA viruses that could be applicable to PPRV. These strategies may contribute to the improvement of existing techniques and/or the development of new reverse genetics systems for PPRV.展开更多
Positive-sense single-stranded RNA(+ssRNA)viruses,the most abundant viruses of eukaryotes in nature,require the synthesis of negative-sense RNA(-RNA)using their genomic(positive-sense)RNA(+RNA)as a template for replic...Positive-sense single-stranded RNA(+ssRNA)viruses,the most abundant viruses of eukaryotes in nature,require the synthesis of negative-sense RNA(-RNA)using their genomic(positive-sense)RNA(+RNA)as a template for replication.Based on current evidence,viral proteins are translated via viral+RNAs,whereas-RNA is considered to be a viral replication intermediate without coding capacity.Here,we report that plant and animal+ssRNA viruses contain small open reading frames(ORFs)in their-RNA(reverse ORFs[rORFs]).Using turnip mosaic virus(TuMV)as a model for plant+ssRNA viruses,we demonstrate that small proteins encoded by rORFs display specific subcellularlocalizations,and confirm the presence of rORF2 in infected cells through mass spectrometry analysis.The protein encoded by TuMV rORF2 forms punctuate granules that are localized in the perinuclear region and co-localized with viral replication complexes.The rORF2 protein can directly interact with the viral RNA-dependent RNA polymerase,and mutation of rORF2 completely abolishes virus infection,whereas ectopic expression of rORF2 rescues the mutant virus.Furthermore,we show that several rORFs in the-RNA of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)have the ability to suppress type l interferon production and facilitate the infection of ve-sicular stomatitis virus.In addition,we provide evidence that TuMV might utilize internal ribosome entry sites to translate these small rORFs.Taken together,these findings indicate that the-RNA of+ssRNA vi-ruses can also have the coding capacity and that small proteins encoded therein play critical roles in viral infection,revealing a viral proteome larger than previously thought.展开更多
Sonchus yellow net virus(SYNV),a negative-sense single-stranded RNA(-ssRNA)virus belonging to the Rhabdoviridae family,encodes six known proteins,including five canonic structural proteins and the nonstructural moveme...Sonchus yellow net virus(SYNV),a negative-sense single-stranded RNA(-ssRNA)virus belonging to the Rhabdoviridae family,encodes six known proteins,including five canonic structural proteins and the nonstructural movement protein Sc4.Using the ORF prediction software ViralORFfinder,we found that SYNV genomic RNA(gRNA)might contain small putative open reading frames(sORFs).However,the functions of these additional sORFs remain largely unexplored.In this study,we identified three conserved sORFs,termed A,B,and C on the gRNA of SYNV.The proteins encoded by these sORFs were found to localize to both the nucleus and cytoplasm.Subcellular localization combined with western blotting revealed a time-dependent decrease in the accumulation of these proteins.Bimolecular fluorescence complementation and co-immunoprecipitation assays confirmed the interactions between A and matrix protein(M),A and phosphoprotein(P),and B and P of SYNV.Mutation of the start codon of B(from AUG to AUA)to block its translation delayed the onset of viral infection,resulting in reduced pathogenicity with milder symptoms and less accumulation of viral RNAs in the late infection stage than the wild-type virus.Ectopic overexpression of A and B via the potato virus X(PVX)-based recombinant vector demonstrated that A and B enhanced PVX-CP accumulation and aggravated mosaic symptoms.These findings reveal the potential coding ability of the gRNA of SYNV and suggest that the identified sORFs may play a significant role in modulating viral pathogenicity.This study also highlights the importance of identifying additional coding capacity in viral genomes,which could provide new insights into the biology of-ssRNA viruses and lead to more opportunities for developing targeted antiviral strategies.展开更多
基金supported by the National Key Research and Development Program of China (2016YFD0500108 and 2016YFE0204100)the International Cooperation Project of CAAS Innovation Program (CAAS-GJHZ201700X)
文摘Peste des petits ruminants(PPR) is a highly contagious transboundary animal disease with a severe socio-economic impact on the livestock industry, particularly in poor countries where it is endemic. Full understanding of PPR virus(PPRV)pathobiology and molecular biology is critical for effective control and eradication of the disease. To achieve these goals,establishment of stable reverse genetics systems for PPRV would play a key role. Unfortunately, this powerful technology remains less accessible and poorly documented for PPRV. In this review, we discussed the current status of PPRV reverse genetics as well as the recent innovations and advances in the reverse genetics of other non-segmented negative-sense RNA viruses that could be applicable to PPRV. These strategies may contribute to the improvement of existing techniques and/or the development of new reverse genetics systems for PPRV.
基金funded by the National Key Research and Development Program of China(2021YFD1400400)to F.L.the National Natural Science Foundation of China(31930089 and 31972244)to X.Z.and F.L.+2 种基金a startup grant for High-level Talents of Fujian Medical University(XRCZX2019019)the Natural Science Foundation of Fujan Province,China(2020J01604)to Q.S.Work in the R.L.-D.lab is partially funded by the ERC-COG grant GemOmics(101044142)to R.L.-D.
文摘Positive-sense single-stranded RNA(+ssRNA)viruses,the most abundant viruses of eukaryotes in nature,require the synthesis of negative-sense RNA(-RNA)using their genomic(positive-sense)RNA(+RNA)as a template for replication.Based on current evidence,viral proteins are translated via viral+RNAs,whereas-RNA is considered to be a viral replication intermediate without coding capacity.Here,we report that plant and animal+ssRNA viruses contain small open reading frames(ORFs)in their-RNA(reverse ORFs[rORFs]).Using turnip mosaic virus(TuMV)as a model for plant+ssRNA viruses,we demonstrate that small proteins encoded by rORFs display specific subcellularlocalizations,and confirm the presence of rORF2 in infected cells through mass spectrometry analysis.The protein encoded by TuMV rORF2 forms punctuate granules that are localized in the perinuclear region and co-localized with viral replication complexes.The rORF2 protein can directly interact with the viral RNA-dependent RNA polymerase,and mutation of rORF2 completely abolishes virus infection,whereas ectopic expression of rORF2 rescues the mutant virus.Furthermore,we show that several rORFs in the-RNA of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)have the ability to suppress type l interferon production and facilitate the infection of ve-sicular stomatitis virus.In addition,we provide evidence that TuMV might utilize internal ribosome entry sites to translate these small rORFs.Taken together,these findings indicate that the-RNA of+ssRNA vi-ruses can also have the coding capacity and that small proteins encoded therein play critical roles in viral infection,revealing a viral proteome larger than previously thought.
基金supported by the National Natural Science Foundation of China(32172385,32202255 and 32320103010)Yunnan Provincial Science and Technology Project(202202AE090022).
文摘Sonchus yellow net virus(SYNV),a negative-sense single-stranded RNA(-ssRNA)virus belonging to the Rhabdoviridae family,encodes six known proteins,including five canonic structural proteins and the nonstructural movement protein Sc4.Using the ORF prediction software ViralORFfinder,we found that SYNV genomic RNA(gRNA)might contain small putative open reading frames(sORFs).However,the functions of these additional sORFs remain largely unexplored.In this study,we identified three conserved sORFs,termed A,B,and C on the gRNA of SYNV.The proteins encoded by these sORFs were found to localize to both the nucleus and cytoplasm.Subcellular localization combined with western blotting revealed a time-dependent decrease in the accumulation of these proteins.Bimolecular fluorescence complementation and co-immunoprecipitation assays confirmed the interactions between A and matrix protein(M),A and phosphoprotein(P),and B and P of SYNV.Mutation of the start codon of B(from AUG to AUA)to block its translation delayed the onset of viral infection,resulting in reduced pathogenicity with milder symptoms and less accumulation of viral RNAs in the late infection stage than the wild-type virus.Ectopic overexpression of A and B via the potato virus X(PVX)-based recombinant vector demonstrated that A and B enhanced PVX-CP accumulation and aggravated mosaic symptoms.These findings reveal the potential coding ability of the gRNA of SYNV and suggest that the identified sORFs may play a significant role in modulating viral pathogenicity.This study also highlights the importance of identifying additional coding capacity in viral genomes,which could provide new insights into the biology of-ssRNA viruses and lead to more opportunities for developing targeted antiviral strategies.
