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Said A Ghabrial - One of the best experts on this subject based on the ideXlab platform.

  • molecular characterization of naturally occurring rna1 recombinants of the Comovirus bean pod mottle virus
    Phytopathology, 2007
    Co-Authors: Chunquan Zhang, Said A Ghabrial
    Abstract:

    ABSTRACT The Bean pod mottle virus, a member of the genus Comovirus, has a bipartite genome consisting of RNA1 and RNA2. We previously reported the occurrence in nature of two distinct subgroups of BPMV strains (subgroups I and II), as well as reassortants between the two subgroups. Here, we report on the isolation and molecular characterization of RNA1 recombinants from soybean plants infected with the partial diploid reassortant strain IL-Cb1, which induces very severe symptoms on soybean. cDNA cloning and sequencing of RNA1 from strain IL-Cb1 revealed the presence of chimeric and mosaic recombinant RNA1s. The full-length mosaic and chimeric recombinant RNA1s were infectious and induced mild symptoms on soybean. Although the recombinant RNA1 accumulated to high levels in the absence of wild-type RNA1, its accumulation level was low in mixed infections with wild-type RNA1. Recombinant RNA1 molecules with similar structures to the naturally occurring recombinant RNA1s were generated in soybean after four ...

  • the bean pod mottle virus proteinase cofactor and putative helicase are symptom severity determinants
    Virology, 2005
    Co-Authors: Said A Ghabrial
    Abstract:

    Abstract 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.

  • soybean mosaic potyvirus enhances the titer of two Comoviruses in dually infected soybean plants
    Phytopathology, 1992
    Co-Authors: J R Anjos, U Jarlfors, Said A Ghabrial
    Abstract:

    Soybean plants dually infected with soybean mosaic potyvirus (SMV) and either one of two Comoviruses, cowpea mosaic virus (CMPV) or bean pod mottle virus (BPMV), showed greatly increased symptom severity over that induced by the individual viruses. The titer of the Comovirus (but not of SMV) in the dually infected plants was significantly higher than that in singly infected plants. Electron microscopic examination of thin sections of leaves from dually infected plants revealed that both SMV and CPMV (or BPMV) were present in the same cells (...)

Joh F Atkins - One of the best experts on this subject based on the ideXlab platform.

  • stimulation of stop codon readthrough frequent presence of an extended 3 rna structural element
    Nucleic Acids Research, 2011
    Co-Authors: Andrew E Firth, Raymond F Gesteland, Norma M Wills, Joh F Atkins
    Abstract:

    In Sindbis, Venezuelan equine encephalitis and related alphaviruses, the polymerase is translated as a fusion with other non-structural proteins via readthrough of a UGA stop codon. Surprisingly, earlier work reported that the signal for efficient readthrough comprises a single cytidine residue 3'-adjacent to the UGA. However, analysis of variability at synonymous sites revealed strikingly enhanced conservation within the ∼ 150 nt 3'-adjacent to the UGA, and RNA folding algorithms revealed the potential for a phylogenetically conserved stem-loop structure in the same region. Mutational analysis of the predicted structure demonstrated that the stem-loop increases readthrough by up to 10-fold. The same computational analysis indicated that similar RNA structures are likely to be relevant to readthrough in certain plant virus genera, notably Furovirus, Pomovirus, Tobravirus, Pecluvirus and Benyvirus, as well as the Drosophilia gene kelch. These results suggest that 3' RNA stimulatory structures feature in a much larger proportion of readthrough cases than previously anticipated, and provide a new criterion for assessing the large number of cellular readthrough candidates that are currently being revealed by comparative sequence analysis.

  • Stimulation of stop codon readthrough: frequent presence of an extended 3′ RNA structural element
    Nucleic acids research, 2011
    Co-Authors: Andrew E Firth, Raymond F Gesteland, Norma M Wills, Joh F Atkins
    Abstract:

    In Sindbis, Venezuelan equine encephalitis and related alphaviruses, the polymerase is translated as a fusion with other non-structural proteins via readthrough of a UGA stop codon. Surprisingly, earlier work reported that the signal for efficient readthrough comprises a single cytidine residue 3'-adjacent to the UGA. However, analysis of variability at synonymous sites revealed strikingly enhanced conservation within the ∼ 150 nt 3'-adjacent to the UGA, and RNA folding algorithms revealed the potential for a phylogenetically conserved stem-loop structure in the same region. Mutational analysis of the predicted structure demonstrated that the stem-loop increases readthrough by up to 10-fold. The same computational analysis indicated that similar RNA structures are likely to be relevant to readthrough in certain plant virus genera, notably Furovirus, Pomovirus, Tobravirus, Pecluvirus and Benyvirus, as well as the Drosophilia gene kelch. These results suggest that 3' RNA stimulatory structures feature in a much larger proportion of readthrough cases than previously anticipated, and provide a new criterion for assessing the large number of cellular readthrough candidates that are currently being revealed by comparative sequence analysis.

  • Stimulation of stop codon readthrough: frequent presence of an extended 39 RNA structural element
    2011
    Co-Authors: Andrew E Firth, Norma M Wills, Raymond F. Gestel, Joh F Atkins
    Abstract:

    In Sindbis, Venezuelan equine encephalitis and related alphaviruses, the polymerase is translated as a fusion with other non-structural proteins via readthrough of a UGA stop codon. Surprisingly, earlier work reported that the signal for efficient readthrough comprises a single cytidine residue 30-adjacent to the UGA. However, analysis of vari-ability at synonymous sites revealed strikingly enhanced conservation within the 150nt 30-adjacent to the UGA, and RNA folding algorithms revealed the potential for a phylogenetically conserved stem–loop structure in the same region. Mutational analysis of the predicted structure demonstrated that the stem–loop increases readthrough by up to 10-fold. The same computa-tional analysis indicated that similar RNA structures are likely to be relevant to readthrough in certain plant virus genera, notably Furovirus, Pomovirus, Tobravirus, Pecluvirus and Benyvirus, as well as the Drosophilia gene kelch. These results suggest that 30 RNA stimulatory structures feature in a much larger proportion of readthrough cases than previously anticipated, and provide a new criterion for assessing the large number of cellular readthrough candidates that are currently being revealed by comparative sequence analysis

George P Lomonossoff - One of the best experts on this subject based on the ideXlab platform.

  • the 24 kda proteinases of Comoviruses are virus specific in cis as well as in trans
    Journal of General Virology, 1996
    Co-Authors: Michael Shanks, Johannes T Dessens, George P Lomonossoff
    Abstract:

    To investigate the specificity of comoviral 24 kDa (‘24K’) proteinases, a full-length cDNA copy of red clover mottle virus (RCMV) RNA 1 has been cloned downstream of a T7 promoter. Translation in rabbit reticulocyte lysates of in vitro transcripts from this clone resulted in the synthesis of a 200K protein which was processed in a manner similar to that of the equivalent protein from cowpea mosaic virus (CPMV). Full-length cDNA clones of the RNA 1 molecules of RCMV and CPMV were used to create hybrid RNA 1 molecules. RNA transcribed in vitro from these hybrids was translated in vitro and the ability of the 24K proteinase from one Comovirus to cleave the 32K/170K processing site from the other assessed. The results of the experiments show that the 24K proteinases are virus-specific in cis.

  • identification and characterization of pseudo recombinants of red clover mottle Comovirus
    Journal of General Virology, 1992
    Co-Authors: P Oxelfelt, Michael Shanks, A K Widmark, George P Lomonossoff
    Abstract:

    cDNA clones specific for the two genomic RNAs of strain O of the Comovirus red clover mottle virus (RCMV) were constructed. Using these clones, in conjunction with clones specific for RNAs of RCMV strain S, local lesion isolates containing reciprocal pseudo-recombinants between strains S and O were identified. Investigation of the biological properties of these pseudo-recombinants showed that the ability of RCMV to infect Chenopodium quinoa is determined by B RNA. The results also suggest that both RNAs are involved in symptom formation in Pisum sativum. Analysis of the strain O clones enabled the sequences at the 3′ ends of both genomic RNAs of strain O to be determined. Comparison of these sequences with the corresponding region of the strain S RNAs suggests that the 3′ terminal sequences critical for replicase recognition may lie somewhat upstream of the poly(A) tract.

  • Comoviruses and enteroviruses share a t cell epitope
    Virology, 1992
    Co-Authors: M A Beck, Beth Ann G Coller, Nora M Chapman, G Hufnagel, Steven Tracy, John E. Johnson, George P Lomonossoff
    Abstract:

    Abstract An in vitro murine T cell proliferation assay was used to determine whether an antigenic epitope(s) recognized by enterovirus-immune T cells is held in common between plant Comoviruses and human enteroviruses. Splenocytes isolated from C3H/HeJ mice infected with coxsackievirus B3 (CVB3) proliferated in vitro not only against a variety of enterovirus (CVB2, CVB3, CVB6, CVA16, PV1) antigens, but against Comovirus (CPMV, BPMV) antigens as well. Splenocytes from mice inoculated with bean pod mottle virus (BPMV) also proliferated in response to comoviral and enteroviral antigens in vitro . However, if the viral inocula were highly purified prior to inoculation, then the splenocyte response was generated only against the group used to inoculate, suggesting that the epitope shared between the Comoviruses and the enteroviruses resided in the nonstructural region. B (nonstructural) and M (structural) genomic segments of CPMV were translated in rabbit reticulocyte lysates and used as in vitro antigens. Splenocytes from mice inoculated with live CVB3 proliferated in response to the B-RNA-encoded but not the M-RNA-encoded polypeptides, confirming the nonstructural coding region location of the common epitope. Comparison of predicted amino acid sequences in the nonstructural coding regions of the Comoviruses and picornaviruses suggested a potentially immunogenic linear epitope in protein 2C. The consensus peptide LEEKGI was synthezized and shown to be immunogenic for both BPMV- and CVB3-immune splenocytes.b

Jari P. T. Valkonen - One of the best experts on this subject based on the ideXlab platform.

  • Complete sequence of RNA 1 and the presence of tRNA-like structures in all RNAs of potato mop-top virus, genus Pomovirus.
    Journal of General Virology, 1999
    Co-Authors: Eugene I. Savenkov, Maria Sandgren, Jari P. T. Valkonen
    Abstract:

    The complete nucleotide sequence (6043 nt) of RNA 1 from Potato mop-top virus (PMTV-Sw), the type member of the genus Pomovirus, was determined. The first (5′-terminal) open reading frame (ORF 1) encodes a predicted protein of 148 kDa. ORF 2 extends through the opal stop codon of ORF 1 producing a predicted readthrough protein of 206 kDa which resembles the RNA-dependent RNA polymerases (RdRp) of other fungal-transmitted viruses. It includes a methyltransferase, a helicase and a GDD RdRp motif, respectively. Phylogenetic analyses of RdRps indicated that PMTV is most closely related to Beet soil-borne virus (genusPomovirus), Broad bean necrosis virus (genus Pomovirus) and Soil-borne wheat mosaic virus (genus Furovirus), and is more distantly related to the other viruses of the former furovirus group. The 5′ and 3′ termini of RNA 1 in PMTV contained untranslated regions (UTR) of 114 nt and 489 nt, respectively. The 3′-UTR of RNA 1 contained a tRNA-like structure, which has previously been reported in the 3′-UTR of RNA 2 but not RNA 3. However, in this study, the tRNA-like structure was also found in the 3′-UTR of RNA 3, which confirms its presence in the 3′-UTRs of all three RNAs of PMTV.

  • Complete sequence of RNA 1 and presence of tRNA-like structures in all RNAs of Potato mop-top pomovirus
    1999
    Co-Authors: Eugene I. Savenkov, Maria S, Jari P. T. Valkonen
    Abstract:

    The complete nucleotide sequence (6043 nt) of RNA 1 from Potato mop-top virus (PMTV-Sw), the type member of the genus Pomovirus, was determined. The first (5«-terminal) open reading frame (ORF 1) encodes a predicted protein of 148 kDa. ORF 2 extends through the opal stop codon of ORF 1 producing a predicted readthrough protein of 206 kDa which resembles the RNA-dependent RNA polymerases (RdRp) of other fungal-transmitted viruses. It includes a methyl-transferase, a helicase and a GDD RdRp motif, respectively. Phylogenetic analyses of RdRps indicated that PMTV is most closely related to Beet soil-borne virus (genus Pomovirus), Broad bean necrosis virus (genus Pomovirus) and Soil-borne wheat mosaic virus (genus Furovirus), and is more distantly related to the other viruses of the former furovirus group. The 5 « and 3 « termini of RNA 1 in PMTV contained untranslated regions (UTR) of 114 nt and 489 nt, respectively. The 3«-UTR of RNA 1 contained a tRNA-like structure, which has previously been reported in the 3«-UTR of RNA 2 but not RNA 3. However, in this study, the tRNA-like structure was also found in the 3«-UTR of RNA 3, which confirms its presence in the 3«-UTRs of all three RNAs of PMTV

Dennis Gonsalves - One of the best experts on this subject based on the ideXlab platform.

  • evidence that resistance in squash mosaic Comovirus coat protein transgenic plants is affected by plant developmental stage and enhanced by combination of transgenes from different lines
    Journal of General Virology, 2000
    Co-Authors: Fuhjyh Jan, Sheng-zhi Pang, David M. Tricoli, Dennis Gonsalves
    Abstract:

    Three transgenic lines of squash hemizygous for the coat protein genes of squash mosaic virus (SqMV) were shown previously to have resistant (SqMV-127), susceptible (SqMV-22) or recovery (SqMV-3) phenotypes. Post-transcriptional gene silencing (PTGS) was the underlying mechanism for resistance of SqMV-127. Here, experiments conducted to determine the mechanism of the recovery phenotype and whether enhanced resistance could be obtained by combining transgenes from susceptible and recovery plants are reported. Upper leaves of SqMV-3 plants were sampled for Northern analysis at 17, 31 and 45 days after germination (DAG) and a proportion of plants were inoculated with SqMV. SqMV-3 plants inoculated at a young stage (17 DAG) showed susceptible or recovery phenotypes. However, a number of plants inoculated at later developmental stages (31 or 45 DAG) were resistant to infection. Resistance of recovery plants was due to PTGS that was activated at a later developmental stage, independent of virus infection. Similar results were observed with plants grown under field conditions. To investigate the interactions of transgenes, progeny of crosses between SqMV-127, -3 and -22 were inoculated with SqMV. Progeny with the transgene of line 127 were resistant. However, a number of plants with transgenes from the recovery and susceptible lines or the self-pollinated recovery line were resistant even when inoculated at a young stage. Northern analysis suggested that resistance was due to PTGS. The results reveal that the timing of PTGS and consequent resistance of the transgenic plants were affected by their developmental stage and the interaction of transgene inserts.

  • Resistance to squash mosaic Comovirus in transgenic squash plants expressing its coat protein genes
    Molecular Breeding, 2000
    Co-Authors: Sheng-zhi Pang, Marc Fuchs, David M. Tricoli, Paul F. Russell, Kim J. Carney, John S. Hu, Hector D. Quemada, Dennis Gonsalves
    Abstract:

    The approach of pathogen-derived resistance was investigated as a means to develop squash mosaic Comovirus (SqMV)-resistant cucurbits. Transgenic squash lines with both coat protein (CP) genes of the melon strain of SqMV were produced and crossed with nontransgenic squash. Further greenhouse, screenhouse and field tests were done with R_1 plants from three independent lines that showed susceptible, recovery, or resistant phenotypes after inoculations with SqMV. Nearly all inoculated plants of the resistant line (SqMV-127) were resistant under greenhouse and field conditions and less so under screenhouse conditions. Plants of the recovery phenotype line (SqMV-3) were susceptible when inoculated at the cotyledon stage but leaves that developed later did not show symptoms. The susceptible line (SqMV-22) developed symptoms that persisted and spread throughout the plant. Plants were also analyzed for transcription rates of the CP transgenes and steady state transgene RNA levels. Results showed that the resistant line SqMV-127 displayed post-transcriptional silencing of the CP transgene as evidenced by high transcription rates but concomitant low accumulation of transgene transcripts. This is the first report on the development of transgenic squash that are resistant to SqMV.