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P. M. Waterhouse - One of the best experts on this subject based on the ideXlab platform.

  • Rice ragged stunt Oryzavirus genome segment S4 could encode an RNA dependent RNA polymerase and a second protein of unknown function
    Archives of Virology, 1998
    Co-Authors: N. M. Upadhyaya, K. Ramm, J. A. Gellatly, W. Kositratana, P. M. Waterhouse
    Abstract:

    The complete nucleotide sequence of genome segment S4 of rice ragged stunt Oryzavirus (RRSV, Thai-isolate) was determined. The 3823 bp sequence contains two large open reading frames (ORFs). ORF1, spanning nucleotides 12 to 3776, is capable of encoding a protein of M _ r 141,380 (P4a). The P4a amino acid sequence predicted from the nucleotide sequence contains sequence motifs conserved in RNA-dependent RNA polymerases (RDRPs). When compared for evolutionary relationships with RDRPs of other reoviruses using the amino acid sequences around the conserved GDD motif, P4a was shown to be more related to Nilaparvata lugens reovirus and reovirus serotype 3 than to rice dwarf phytoreovirus, bovine rotavirus or bluetongue virus. The ORF2, spanning nucleotides 491 to 1468, is out of frame with ORF1 and is capable of encoding a protein of 36, 920 (P4b). Coupled in vitro transcription-translation from cloned ORF2 in wheat germ extract confirmed he existence of ORF2 but in vivo production and possible function of P4b is yet to be determined.

  • Rice ragged stunt Oryzavirus genome segments S7 and S10 encode non-structural proteins of M_r 68 025 (Pns7) and M_r 32 364 (Pns10)
    Archives of Virology, 1997
    Co-Authors: N. M. Upadhyaya, K. Ramm, J. A. Gellatly, W. Kositratana, P. M. Waterhouse
    Abstract:

    The nucleotide sequences of genome segments S7 and S10 of a Thai-isolate of rice ragged stunt virus (RRSV) were determined. The 1 938 bp S7 sequence contains a single large open reading frame (ORF) spanning nucleotides 20 to 1 843 that is predicted to encode a protein of M _ r 68 025. The 1 162 bp S10 sequence has a major ORF spanning nucleotides 142 to 1 032 that is predicted to encode a protein of M _ r 32 364. This S10 ORF is preceded by a small ORF (nt 20–55) which is probably a minicistron. Coupled in vitro transcription-translation from the two major ORFs gave protein products of the expected sizes. However, no protein was visualised from S10 when the small ORF sequence was included. Proteins were expressed in Escherichia coli from the full length ORF of S7 (P7) and from a segment of the S10 ORF (P10) fused to the ORF of glutathione S-transferase (GST). Neither fusion protein was recognised by polyclonal antibodies raised against RRSV particles. Furthermore, polyclonal antibodies raised against GST-P7 fusion protein did not recognise any virion structural polypeptides. These data strongly suggest that the proteins P7 and P10 do not form part of RRSV particle. This is further supported by observed sequence homology (though very weak) of predicted RRSV P7 and P10 with those of rice dwarf virus (RDV) non-structural proteins Pns6 and Pns9, respectively.

  • TheM _ r 43K major capsid protein of rice ragged stunt Oryzavirus is a post-translationally processed product of aM _ r 67 348 polypeptide encoded by genome segment 8
    Archives of Virology, 1996
    Co-Authors: N. M. Upadhyaya, W. Kositratana, E. Zinkowsky, P. M. Waterhouse
    Abstract:

    The nucleotide sequence of DNA complementary to rice ragged stunt Oryzavirus (RRSV) genome segment 8 (S8) of an isolate from Thailand was determined. RRSV S8 is 1914 bp in size and contains a single large open reading frame (ORF) spanning nucleotides 23 to 1810 which is capable of encoding a protein of M _ r 67 348. The N-terminal amino acid sequence of a ∼43K virion polypeptide matched to that inferred for an internal region of the S8 coding sequence. These data suggest that the 43K protein is encoded by S8 and is derived by a proteolytic cleavage. Predicted polypeptide sizes from this possible cleavage of S8 protein are 26K and 42K. Polyclonal antibodies raised against a maltose binding protein (MBP)-S8 fusion polypeptide (expressed in Escherichia coli ) recognised four RRSV particle associated polypeptides of M_ r 67K, 46K, 43K and 26K and all except the 26K polypeptide were also highly immunoreactive to polyclonal antibodies raised against purified RRSV particles. Cleavage of the MBP-S8 fusion polypeptide with protease Factor X produced the expected 40K MBP and two polypeptides of apparent M _ r 46K and 26K. Antibodies to purified RRSV particles reacted strongly with the intact fusion protein and the 46K cleavage product but weakly to the 26K product. Furthermore, in vitro transcription and translation of the S8 coding region revealed a post-translational self cleavage of the 67K polypeptide to 46K and 26K products. These data indicate that S8 encodes a structural polypeptide, the majority of which is auto-catalytically cleaved to 26K and 46K proteins. The data also suggest that the 26K protein is the self cleaving protease and that the 46K product is further processed or undergoes stable conformational changes to a ∼43K major capsid protein.

  • Molecular analysis of rice ragged stunt Oryzavirus segment 9 and sequence conservation among isolates from Thailand and India
    Archives of Virology, 1995
    Co-Authors: N. M. Upadhyaya, W. Kositratana, M. Yang, A. Ghosh, P. M. Waterhouse
    Abstract:

    Nucleotide sequences of rice ragged stunt virus (RRSV) genome segment 9 (S9) from a Thai and an Indian isolate were determined. Both sequences are 1132 bp long, contain a single large open reading frame (ORF) spanning nucleotide residues 14 to 1027 and are capable of encoding a protein of 38.5K. The two isolates are 94.6% and 99.4% identical at the nucleotide and amino acid level, respectively. The authenticity of these coding sequences was confirmed by identifying a ∼38K protein in the RRSV particle with an N-terminal amino acid sequence identical to that inferred from the S9 ORF. Furthermore, cDNA of S9 from each isolate incorporated into the bacterial expression vector pGEX3-X produced a fusion protein that reacted with antibodies raised against purified RRSV particles. Cleaving these fusion proteins with protease factor X liberated a ∼38K polypeptide.

Guohui Zhou - One of the best experts on this subject based on the ideXlab platform.

  • southern rice black streaked dwarf virus alters insect vectors host orientation preferences to enhance spread and increase rice ragged stunt virus co infection
    Phytopathology, 2014
    Co-Authors: Han Wang, Donglin Xu, Lingling Pu, Guohui Zhou
    Abstract:

    In recent years, Southern rice black-streaked dwarf virus (SRBSDV), a tentative species in the genus Fijivirus (family Reoviridae), has spread rapidly and caused serious rice losses in eastern and southeastern Asia. With this virus spread, Rice ragged stunt virus (RRSV, genus Oryzavirus, family Reoviridae) became more common in southern China, usually in co-infection with the former. SRBSDV and RRSV are transmitted by two different species of planthoppers, white-backed planthopper (WBPH, Sogatella furcifera) and brown planthopper (BPH, Nilaparvata lugens), respectively, in a persistent, circulative, propagative manner. In this study, using a Y-shape olfactometer-based device, we tested the host preference of three types of macropterous WBPH adults for healthy or SRBSDV-infected rice plants. The results showed that virus-free WBPHs significantly preferred infected rice plants to healthy plants, whereas both the viruliferous and nonviruliferous WBPHs preferred healthy plants to infected plants. In additional tests, we found that the BPHs significantly preferred healthy plants when they were virus free, whereas RRSV-carrying BPHs preferred SRBSDV-infected rice plants. From these findings, we propose that plant viruses may alter host selection preference of vectors to enhance their spread and that of insects vectoring another virus to result in co-infection with more than one virus.

  • Synergism between southern rice black-streaked dwarf virus and rice ragged stunt virus enhances their insect vector acquisition.
    Phytopathology, 2014
    Co-Authors: Han Wang, Guohui Zhou
    Abstract:

    ABSTRACT Southern rice black-streaked dwarf virus (SRBSDV), a tentative species in the genus Fijivirus, family Reoviridae, is a novel rice virus transmitted by the white-backed planthopper (Sogatella furcifera). Since its discovery in 2001, SRBSDV has spread rapidly throughout eastern and southeastern Asia and caused large rice losses in China and Vietnam. Rice ragged stunt virus (RRSV) (genus Oryzavirus, family Reoviridae) is a common rice virus vectored by the brown planthopper (Nilaparvata lugens). RRSV is also widely distributed in eastern and southeastern Asia but has not previously caused serious problems in China owing to its low incidence. With SRBSDV's spread, however, RRSV has become increasingly common in China, and is frequently found in co-infection with SRBSDV. In this study, we show that SRBSDV and RRSV interact synergistically, the first example of synergism between plant viruses in the family Reoviridae. Rice plants co-infected with both viruses displayed enhanced stunting, earlier sympto...

W. Kositratana - One of the best experts on this subject based on the ideXlab platform.

  • Rice ragged stunt Oryzavirus genome segment S4 could encode an RNA dependent RNA polymerase and a second protein of unknown function
    Archives of Virology, 1998
    Co-Authors: N. M. Upadhyaya, K. Ramm, J. A. Gellatly, W. Kositratana, P. M. Waterhouse
    Abstract:

    The complete nucleotide sequence of genome segment S4 of rice ragged stunt Oryzavirus (RRSV, Thai-isolate) was determined. The 3823 bp sequence contains two large open reading frames (ORFs). ORF1, spanning nucleotides 12 to 3776, is capable of encoding a protein of M _ r 141,380 (P4a). The P4a amino acid sequence predicted from the nucleotide sequence contains sequence motifs conserved in RNA-dependent RNA polymerases (RDRPs). When compared for evolutionary relationships with RDRPs of other reoviruses using the amino acid sequences around the conserved GDD motif, P4a was shown to be more related to Nilaparvata lugens reovirus and reovirus serotype 3 than to rice dwarf phytoreovirus, bovine rotavirus or bluetongue virus. The ORF2, spanning nucleotides 491 to 1468, is out of frame with ORF1 and is capable of encoding a protein of 36, 920 (P4b). Coupled in vitro transcription-translation from cloned ORF2 in wheat germ extract confirmed he existence of ORF2 but in vivo production and possible function of P4b is yet to be determined.

  • Rice ragged stunt Oryzavirus genome segments S7 and S10 encode non-structural proteins of M_r 68 025 (Pns7) and M_r 32 364 (Pns10)
    Archives of Virology, 1997
    Co-Authors: N. M. Upadhyaya, K. Ramm, J. A. Gellatly, W. Kositratana, P. M. Waterhouse
    Abstract:

    The nucleotide sequences of genome segments S7 and S10 of a Thai-isolate of rice ragged stunt virus (RRSV) were determined. The 1 938 bp S7 sequence contains a single large open reading frame (ORF) spanning nucleotides 20 to 1 843 that is predicted to encode a protein of M _ r 68 025. The 1 162 bp S10 sequence has a major ORF spanning nucleotides 142 to 1 032 that is predicted to encode a protein of M _ r 32 364. This S10 ORF is preceded by a small ORF (nt 20–55) which is probably a minicistron. Coupled in vitro transcription-translation from the two major ORFs gave protein products of the expected sizes. However, no protein was visualised from S10 when the small ORF sequence was included. Proteins were expressed in Escherichia coli from the full length ORF of S7 (P7) and from a segment of the S10 ORF (P10) fused to the ORF of glutathione S-transferase (GST). Neither fusion protein was recognised by polyclonal antibodies raised against RRSV particles. Furthermore, polyclonal antibodies raised against GST-P7 fusion protein did not recognise any virion structural polypeptides. These data strongly suggest that the proteins P7 and P10 do not form part of RRSV particle. This is further supported by observed sequence homology (though very weak) of predicted RRSV P7 and P10 with those of rice dwarf virus (RDV) non-structural proteins Pns6 and Pns9, respectively.

  • Rice ragged stunt Oryzavirus genome segments S7 and S10 encode non-structural proteins of M(r) 68,025 (Pns7) and M(r) 32,364 (Pns10).
    Archives of virology, 1997
    Co-Authors: Narayana M Upadhyaya, K. Ramm, J. A. Gellatly, W. Kositratana, Peter M Waterhouse
    Abstract:

    The nucleotide sequences of genome segments S7 and S10 of a Thai-isolate of rice ragged stunt virus (RRSV) were determined. The 1 938 bp S7 sequence contains a single large open reading frame (ORF) spanning nucleotides 20 to 1 843 that is predicted to encode a protein of M r 68 025. The 1 162 bp S10 sequence has a major ORF spanning nucleotides 142 to 1 032 that is predicted to encode a protein of M r 32 364. This S10 ORF is preceded by a small ORF (nt 20–55) which is probably a minicistron. Coupled in vitro transcription-translation from the two major ORFs gave protein products of the expected sizes. However, no protein was visualised from S10 when the small ORF sequence was included. Proteins were expressed in Escherichia coli from the full length ORF of S7 (P7) and from a segment of the S10 ORF (P10) fused to the ORF of glutathione S-transferase (GST). Neither fusion protein was recognised by polyclonal antibodies raised against RRSV particles. Furthermore, polyclonal antibodies raised against GST-P7 fusion protein did not recognise any virion structural polypeptides. These data strongly suggest that the proteins P7 and P10 do not form part of RRSV particle. This is further supported by observed sequence homology (though very weak) of predicted RRSV P7 and P10 with those of rice dwarf virus (RDV) non-structural proteins Pns6 and Pns9, respectively.

  • Rice ragged stunt Oryzavirus genome segments S7 and S10 encode non-structural proteins of M(r) 68 025 (Pns7) and M(r) 32364 (Pns10) : brief report
    Archives of Virology, 1997
    Co-Authors: Narayana M Upadhyaya, K. Ramm, J. A. Gellatly, W. Kositratana, Peter M Waterhouse
    Abstract:

    The nucleotide sequences of genome segments S7 and S10 of a Thai-isolate of rice ragged stunt virus (RRSV) were determined. The 1938 bp S7 sequence contains a single large open reading frame (ORF) spanning nucleotides 20 to 1 843 that is predicted to encode a protein of M(r) 68 025. The 1 162 bp S10 sequence has a major ORF spanning nucleotides 142 to 1 032 that is predicted to encode a protein of M(r) 32364. This S10 ORF is preceded by a small ORF (nt 20-55) which is probably a minicistron. Coupled in vitro transcription-translation from the two major ORFs gave protein products of the expected sizes. However, no protein was visualised from S10 when the small ORF sequence was included. Proteins were expressed in Escherichia coli from the full length ORF of S7 (P7) and from a segment of the S10 ORF (P10) fused to the ORF of glutathione S-transferase (GST). Neither fusion protein was recognised by polyclonal antibodies raised against RRSV particles. Furthermore, polyclonal antibodies raised against GST-P7 fusion protein did not recognise any virion structural polypeptides. These data strongly suggest that the proteins P7 and P10 do not form part of RRSV particle. This is further supported by observed sequence homology (though very weak) of predicted.

  • TheM _ r 43K major capsid protein of rice ragged stunt Oryzavirus is a post-translationally processed product of aM _ r 67 348 polypeptide encoded by genome segment 8
    Archives of Virology, 1996
    Co-Authors: N. M. Upadhyaya, W. Kositratana, E. Zinkowsky, P. M. Waterhouse
    Abstract:

    The nucleotide sequence of DNA complementary to rice ragged stunt Oryzavirus (RRSV) genome segment 8 (S8) of an isolate from Thailand was determined. RRSV S8 is 1914 bp in size and contains a single large open reading frame (ORF) spanning nucleotides 23 to 1810 which is capable of encoding a protein of M _ r 67 348. The N-terminal amino acid sequence of a ∼43K virion polypeptide matched to that inferred for an internal region of the S8 coding sequence. These data suggest that the 43K protein is encoded by S8 and is derived by a proteolytic cleavage. Predicted polypeptide sizes from this possible cleavage of S8 protein are 26K and 42K. Polyclonal antibodies raised against a maltose binding protein (MBP)-S8 fusion polypeptide (expressed in Escherichia coli ) recognised four RRSV particle associated polypeptides of M_ r 67K, 46K, 43K and 26K and all except the 26K polypeptide were also highly immunoreactive to polyclonal antibodies raised against purified RRSV particles. Cleavage of the MBP-S8 fusion polypeptide with protease Factor X produced the expected 40K MBP and two polypeptides of apparent M _ r 46K and 26K. Antibodies to purified RRSV particles reacted strongly with the intact fusion protein and the 46K cleavage product but weakly to the 26K product. Furthermore, in vitro transcription and translation of the S8 coding region revealed a post-translational self cleavage of the 67K polypeptide to 46K and 26K products. These data indicate that S8 encodes a structural polypeptide, the majority of which is auto-catalytically cleaved to 26K and 46K proteins. The data also suggest that the 26K protein is the self cleaving protease and that the 46K product is further processed or undergoes stable conformational changes to a ∼43K major capsid protein.

N. M. Upadhyaya - One of the best experts on this subject based on the ideXlab platform.

  • Rice ragged stunt Oryzavirus genome segment S4 could encode an RNA dependent RNA polymerase and a second protein of unknown function
    Archives of Virology, 1998
    Co-Authors: N. M. Upadhyaya, K. Ramm, J. A. Gellatly, W. Kositratana, P. M. Waterhouse
    Abstract:

    The complete nucleotide sequence of genome segment S4 of rice ragged stunt Oryzavirus (RRSV, Thai-isolate) was determined. The 3823 bp sequence contains two large open reading frames (ORFs). ORF1, spanning nucleotides 12 to 3776, is capable of encoding a protein of M _ r 141,380 (P4a). The P4a amino acid sequence predicted from the nucleotide sequence contains sequence motifs conserved in RNA-dependent RNA polymerases (RDRPs). When compared for evolutionary relationships with RDRPs of other reoviruses using the amino acid sequences around the conserved GDD motif, P4a was shown to be more related to Nilaparvata lugens reovirus and reovirus serotype 3 than to rice dwarf phytoreovirus, bovine rotavirus or bluetongue virus. The ORF2, spanning nucleotides 491 to 1468, is out of frame with ORF1 and is capable of encoding a protein of 36, 920 (P4b). Coupled in vitro transcription-translation from cloned ORF2 in wheat germ extract confirmed he existence of ORF2 but in vivo production and possible function of P4b is yet to be determined.

  • Rice ragged stunt Oryzavirus genome segments S7 and S10 encode non-structural proteins of M_r 68 025 (Pns7) and M_r 32 364 (Pns10)
    Archives of Virology, 1997
    Co-Authors: N. M. Upadhyaya, K. Ramm, J. A. Gellatly, W. Kositratana, P. M. Waterhouse
    Abstract:

    The nucleotide sequences of genome segments S7 and S10 of a Thai-isolate of rice ragged stunt virus (RRSV) were determined. The 1 938 bp S7 sequence contains a single large open reading frame (ORF) spanning nucleotides 20 to 1 843 that is predicted to encode a protein of M _ r 68 025. The 1 162 bp S10 sequence has a major ORF spanning nucleotides 142 to 1 032 that is predicted to encode a protein of M _ r 32 364. This S10 ORF is preceded by a small ORF (nt 20–55) which is probably a minicistron. Coupled in vitro transcription-translation from the two major ORFs gave protein products of the expected sizes. However, no protein was visualised from S10 when the small ORF sequence was included. Proteins were expressed in Escherichia coli from the full length ORF of S7 (P7) and from a segment of the S10 ORF (P10) fused to the ORF of glutathione S-transferase (GST). Neither fusion protein was recognised by polyclonal antibodies raised against RRSV particles. Furthermore, polyclonal antibodies raised against GST-P7 fusion protein did not recognise any virion structural polypeptides. These data strongly suggest that the proteins P7 and P10 do not form part of RRSV particle. This is further supported by observed sequence homology (though very weak) of predicted RRSV P7 and P10 with those of rice dwarf virus (RDV) non-structural proteins Pns6 and Pns9, respectively.

  • TheM _ r 43K major capsid protein of rice ragged stunt Oryzavirus is a post-translationally processed product of aM _ r 67 348 polypeptide encoded by genome segment 8
    Archives of Virology, 1996
    Co-Authors: N. M. Upadhyaya, W. Kositratana, E. Zinkowsky, P. M. Waterhouse
    Abstract:

    The nucleotide sequence of DNA complementary to rice ragged stunt Oryzavirus (RRSV) genome segment 8 (S8) of an isolate from Thailand was determined. RRSV S8 is 1914 bp in size and contains a single large open reading frame (ORF) spanning nucleotides 23 to 1810 which is capable of encoding a protein of M _ r 67 348. The N-terminal amino acid sequence of a ∼43K virion polypeptide matched to that inferred for an internal region of the S8 coding sequence. These data suggest that the 43K protein is encoded by S8 and is derived by a proteolytic cleavage. Predicted polypeptide sizes from this possible cleavage of S8 protein are 26K and 42K. Polyclonal antibodies raised against a maltose binding protein (MBP)-S8 fusion polypeptide (expressed in Escherichia coli ) recognised four RRSV particle associated polypeptides of M_ r 67K, 46K, 43K and 26K and all except the 26K polypeptide were also highly immunoreactive to polyclonal antibodies raised against purified RRSV particles. Cleavage of the MBP-S8 fusion polypeptide with protease Factor X produced the expected 40K MBP and two polypeptides of apparent M _ r 46K and 26K. Antibodies to purified RRSV particles reacted strongly with the intact fusion protein and the 46K cleavage product but weakly to the 26K product. Furthermore, in vitro transcription and translation of the S8 coding region revealed a post-translational self cleavage of the 67K polypeptide to 46K and 26K products. These data indicate that S8 encodes a structural polypeptide, the majority of which is auto-catalytically cleaved to 26K and 46K proteins. The data also suggest that the 26K protein is the self cleaving protease and that the 46K product is further processed or undergoes stable conformational changes to a ∼43K major capsid protein.

  • Molecular analysis of rice ragged stunt Oryzavirus segment 9 and sequence conservation among isolates from Thailand and India
    Archives of Virology, 1995
    Co-Authors: N. M. Upadhyaya, W. Kositratana, M. Yang, A. Ghosh, P. M. Waterhouse
    Abstract:

    Nucleotide sequences of rice ragged stunt virus (RRSV) genome segment 9 (S9) from a Thai and an Indian isolate were determined. Both sequences are 1132 bp long, contain a single large open reading frame (ORF) spanning nucleotide residues 14 to 1027 and are capable of encoding a protein of 38.5K. The two isolates are 94.6% and 99.4% identical at the nucleotide and amino acid level, respectively. The authenticity of these coding sequences was confirmed by identifying a ∼38K protein in the RRSV particle with an N-terminal amino acid sequence identical to that inferred from the S9 ORF. Furthermore, cDNA of S9 from each isolate incorporated into the bacterial expression vector pGEX3-X produced a fusion protein that reacted with antibodies raised against purified RRSV particles. Cleaving these fusion proteins with protease factor X liberated a ∼38K polypeptide.

Peter M Waterhouse - One of the best experts on this subject based on the ideXlab platform.

  • ectopic expression of the spike protein of rice ragged stunt Oryzavirus in transgenic rice plants inhibits transmission of the virus to insects
    Molecular Breeding, 2003
    Co-Authors: Shao Chaogang, Wu Jianhua, Zhou Guoying, Sun Gang, Peng Baozhen, Lei Juanli, Jin Dendi, Chen Shenxiang, Narayana M Upadhyaya, Peter M Waterhouse
    Abstract:

    Rice ragged stunt Oryzavirus (RRSV) replicates in both its insect vector, Nilaparvata lugens, and its plant host, rice, and has a complex multi-component particle bearing spikes on its outer surface. Transgenic rice lines expressing the 39 kDa spike protein showed good resistance to infection by RRSV. Furthermore, N. lugens fed on these plants prior to feeding on RRSV-infected plants were significantly protected against RRSV infection. The viral titre in insects initially fed on transgenic plants and then on RRSV-infected plants was inversely proportional to the levels of the 39 kDa protein expressed in the transgenic plants. This suggests that the 39 kDa protein interferes with the interaction between the intact virus particles and insect cell receptors and that the spike protein of RRSV contributes to vector specificity. This approach would probably be a more environment-friendly and sustainable method of virus control than by actual eradication of insect vectors.

  • Rice ragged stunt Oryzavirus genome segments S7 and S10 encode non-structural proteins of M(r) 68,025 (Pns7) and M(r) 32,364 (Pns10).
    Archives of virology, 1997
    Co-Authors: Narayana M Upadhyaya, K. Ramm, J. A. Gellatly, W. Kositratana, Peter M Waterhouse
    Abstract:

    The nucleotide sequences of genome segments S7 and S10 of a Thai-isolate of rice ragged stunt virus (RRSV) were determined. The 1 938 bp S7 sequence contains a single large open reading frame (ORF) spanning nucleotides 20 to 1 843 that is predicted to encode a protein of M r 68 025. The 1 162 bp S10 sequence has a major ORF spanning nucleotides 142 to 1 032 that is predicted to encode a protein of M r 32 364. This S10 ORF is preceded by a small ORF (nt 20–55) which is probably a minicistron. Coupled in vitro transcription-translation from the two major ORFs gave protein products of the expected sizes. However, no protein was visualised from S10 when the small ORF sequence was included. Proteins were expressed in Escherichia coli from the full length ORF of S7 (P7) and from a segment of the S10 ORF (P10) fused to the ORF of glutathione S-transferase (GST). Neither fusion protein was recognised by polyclonal antibodies raised against RRSV particles. Furthermore, polyclonal antibodies raised against GST-P7 fusion protein did not recognise any virion structural polypeptides. These data strongly suggest that the proteins P7 and P10 do not form part of RRSV particle. This is further supported by observed sequence homology (though very weak) of predicted RRSV P7 and P10 with those of rice dwarf virus (RDV) non-structural proteins Pns6 and Pns9, respectively.

  • Rice ragged stunt Oryzavirus genome segments S7 and S10 encode non-structural proteins of M(r) 68 025 (Pns7) and M(r) 32364 (Pns10) : brief report
    Archives of Virology, 1997
    Co-Authors: Narayana M Upadhyaya, K. Ramm, J. A. Gellatly, W. Kositratana, Peter M Waterhouse
    Abstract:

    The nucleotide sequences of genome segments S7 and S10 of a Thai-isolate of rice ragged stunt virus (RRSV) were determined. The 1938 bp S7 sequence contains a single large open reading frame (ORF) spanning nucleotides 20 to 1 843 that is predicted to encode a protein of M(r) 68 025. The 1 162 bp S10 sequence has a major ORF spanning nucleotides 142 to 1 032 that is predicted to encode a protein of M(r) 32364. This S10 ORF is preceded by a small ORF (nt 20-55) which is probably a minicistron. Coupled in vitro transcription-translation from the two major ORFs gave protein products of the expected sizes. However, no protein was visualised from S10 when the small ORF sequence was included. Proteins were expressed in Escherichia coli from the full length ORF of S7 (P7) and from a segment of the S10 ORF (P10) fused to the ORF of glutathione S-transferase (GST). Neither fusion protein was recognised by polyclonal antibodies raised against RRSV particles. Furthermore, polyclonal antibodies raised against GST-P7 fusion protein did not recognise any virion structural polypeptides. These data strongly suggest that the proteins P7 and P10 do not form part of RRSV particle. This is further supported by observed sequence homology (though very weak) of predicted.

  • The M r 43K major capsid protein of rice ragged stunt Oryzavirus is a post-translationally processed product of a M r 67 348 polypeptide encoded by genome segment 8
    Archives of virology, 1996
    Co-Authors: Narayana M Upadhyaya, W. Kositratana, E. Zinkowsky, Peter M Waterhouse
    Abstract:

    The nucleotide sequence of DNA complementary to rice ragged stunt Oryzavirus (RRSV) genome segment 8 (S8) of an isolate from Thailand was determined. RRSV S8 is 1914 bp in size and contains a single large open reading frame (ORF) spanning nucleotides 23 to 1810 which is capable of encoding a protein ofMr 67 348. The N-terminal amino acid sequence of a ∼43K virion polypeptide matched to that inferred for an internal region of the S8 coding sequence. These data suggest that the 43K protein is encoded by S8 and is derived by a proteolytic cleavage. Predicted polypeptide sizes from this possible cleavage of S8 protein are 26K and 42K. Polyclonal antibodies raised against a maltose binding protein (MBP)-S8 fusion polypeptide (expressed inEscherichia coli) recognised four RRSV particle associated polypeptides of Mr 67K, 46K, 43K and 26K and all except the 26K polypeptide were also highly immunoreactive to polyclonal antibodies raised against purified RRSV particles. Cleavage of the MBP-S8 fusion polypeptide with protease Factor X produced the expected 40K MBP and two polypeptides of apparentMr 46K and 26K. Antibodies to purified RRSV particles reacted strongly with the intact fusion protein and the 46K cleavage product but weakly to the 26K product. Furthermore, in vitro transcription and translation of the S8 coding region revealed a post-translational self cleavage of the 67K polypeptide to 46K and 26K products. These data indicate that S8 encodes a structural polypeptide, the majority of which is auto-catalytically cleaved to 26K and 46K proteins. The data also suggest that the 26K protein is the self cleaving protease and that the 46K product is further processed or undergoes stable conformational changes to a ∼43K major capsid protein.