| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
1 Scottish Crop Research Institute, Invergowrie, Dundee DD2 5DA, UK
2 National Horticultural Research Institute, Rural Development Administration, 475 Imok-Dong, Suwon 440-310, Korea
Correspondence
Peter Palukaitis
Peter.Palukaitis{at}scri.ac.uk
 |
ABSTRACT |
|---|
 TOP ABSTRACT INTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
|---|
Supplementary figures are available in JGV Online.
|
INTRODUCTION |
|---|
|
TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
|---|
; Prins, 2003). However, the use of transgenic plants expressing plant viral sequences has raised a number of environmental concerns regarding various interactions. These include trans-encapsidation (when viral capsid protein genes were employed), synergy of disease between the transgene product and a non-target virus (i.e. a virus unrelated to the viral source of the transgene) and recombination between the transgene mRNA and non-target viruses (Palukaitis, 1991; Palukaitis & Kaplan, 1997; Tepfer, 2002). Recombination has been observed between expressed viral transgenes and the same viruses used as the source of the transgene (Adair & Kearney, 2000; Borja et al., 1999; Greene & Allison, 1994, 1996; Schoelz & Wintermantel, 1993; Varrelmann et al., 2000; Wintermantel & Schoelz, 1996), as well as between two related viruses (i.e. in the same genus) co-infecting plants (Aaziz & Tepfer, 1999; Masuta et al., 1998; White & Morris, 1994). Recombination has also been used to explain the presence of plant viral sequences in plant genomes (Bejarano et al., 1996; Harper et al., 1999; Jakowitsch et al., 1999; Tanne & Sela, 2005). Similarly, sequence analyses indicate that recombination had occurred between closely related viruses (Chare & Holmes, 2006), especially potyviruses (Bousalem et al., 2000; Cervera et al., 1993; Desbiez & Lecoq, 2004; Krause-Sakate et al., 2004; Revers et al., 1996; Tan et al., 2004), in some cases altering the virulence of the virus (Glais et al., 2002; Larsen et al., 2005; Zhong et al., 2005). However, to date, no recombination has been demonstrated experimentally between completely unrelated viruses (i.e. in different families), or between a viral-derived transgene and an unrelated virus. Concerns have been raised concerning possible consequences of such recombination events, giving rise to novel viruses with potentially altered host range (reviewed by Tepfer, 2002). Given the precision of recombination required to generate such novel viruses (i.e. integration into a region of the virus genome that would allow expression of the donor gene without affecting either the replication of the acceptor virus or expression of the other genes in the acceptor virus), it may be rather difficult to detect such recombinant viruses. Therefore, as an approach to examine the consequences of recombination between a virus and a transgene derived from an unrelated virus, three hybrid viruses were generated in vitro, using viral genes previously used as transgenes and plant viruses previously utilized as gene expression vectors. These hybrid viruses were then assessed for their stability in two host species. |
METHODS |
|---|
|
TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
|---|
) were also inoculated with TRV-NIb. The TRV inocula consisted of a mixture of transcript derived from cDNA clones of RNA2 of either wild-type (wt) TRV or TRV-NIb mixed with total RNAs from plants previously infected with TRV RNA1, as described by Mueller et al. (1997). The other inocula consisted of either RNA transcripts obtained by in vitro transcription of the PVX derivative plasmids, or buffered extracts from plants infected with transcripts 3 days post-inoculation (p.i.), for experiments involving TRV and TRV-NIb (Canto & Palukaitis, 2001, 2002). The various in vitro transcripts were prepared using an mMESSAGE mMACHINE T7 transcription kit (Ambion) as previously described (Canto & Palukaitis, 2001; Canto et al., 2004).
Plasmids and constructs
Generation of TRV-NIb.
The PVY sequence containing the NIb ORF (nt 7010–8566) was amplified by PCR from a full-length cDNA clone of a PVYO isolate obtained from the Scottish Crop Research Institute collection (unpublished data). The upstream PCR primer (5'-CATGTTAACATGGCTAAGCACTCTGCGTGG-3') contained an HpaI site (underlined) and an added ATG initiation codon (bold). The downstream PCR primer (5'-CATTCTAGATTGATGGTGTACTTCATAAG-3') contained an XbaI site (underlined). The PCR fragment was digested with HpaI and XbaI and was ligated into the vector TRV-AtALY2-GFP (Uhrig et al., 2004), previously cut with the same two restriction enzymes. The resulting clone, TRV-NIb/GFP, expressed the NIb gene as a fusion to the gene encoding the green fluorescent protein (GFP). To remove the GFP-encoding sequence, the NIb gene was amplified again by PCR using the above upstream primer and 5'-CATCTCGAGGTACCTATTGATGGTGTACTTCATAA-3' as the downstream primer, containing an added termination codon (bold) and a KpnI site (underlined). This PCR fragment was digested with SacI (internal to the NIb sequence) and KpnI, and ligated into TRV-NIb/GFP previously digested with the same enzymes, generating the construct TRV-NIb.
Generation of PVX-NIb.
The NIb gene was amplified from TRV-NIb/GFP by PCR using 5'-ATGCGGCCGATGGCTAAGCACTCTGCGTGG-3' as the upstream primer, containing an EagI site (underlined) and an added ATG codon (bold), and 5'-CATGATATCATTGATGGTGTACTTCATAAGAG-3' as the downstream primer, containing an added termination codon (bold) and an EcoRV site (underlined). The PCR fragment was digested with EagI and EcoRV and was ligated into the PVX vector pTX.P3C2 402 (Baulcombe et al., 1995) previously digested with the same restriction enzymes, generating the construct PVX-NIb.
Generation of PVX-HCPro.
The PVY sequence containing the HCPro ORF (nt 965–2407) was amplified by PCR from the above full-length cDNA clone of PVYO. The upstream PCR primer (5'-ATCCGGCCGATGTCGAATGCTGACAATTTTTGG-3') contained an EagI site (underlined) and an added ATG starting codon (bold). The downstream primer (5'-AGCTGCAGCCCGGGTTAACCAACTCTATAATG-3') contained an added stop codon (bold) and a PstI site (underlined). The PCR fragment was digested with EagI and PstI and was ligated into the PVX vector pTX.P3C2 402 previously digested with EagI and NsiI, to generate the construct PVX-HCPro.
Preparation and analysis of plant nucleic acid samples.
Total RNAs from either mock-inoculated or virus-infected plants were isolated using a buffer–phenol/chloroform extraction method (Canto & Palukaitis, 1998). RNAs were fractionated by agarose gel electrophoresis, blotted to nitrocellulose membranes and hybridized with the specified digoxigenin-labelled RNA probes for TRV RNA2 and PVX RNA, all as described previously (Canto & Palukaitis, 2001; Canto et al., 2004). The TRV probe was specific to the sequences encoding the capsid protein (CP) in TRV RNA2, while the PVX probe was specific to the 3'-terminal sequences.
The total plant RNAs extracted were used for RT-PCR with primers TRV-3' (5'-CGAGAATGTCAATCTCGTAGG-3') or PVX-3' (5'-TGTACTAAAGAAATCCCCATCC-3') for the RT step in a 13 µl reaction done at 37 °C for 60 min. The PCR (30 µl) was done using 5 µl RT reaction products and either the primers TRV-5' (5'-CTGGGAGATGATACGCTG-3') plus TRV-3' or the primers PVX-5' (5'-ATCACAGTGTTGGCTTGC-3') plus PVX-3' in a reaction mixture recommended by the supplier of the Taq DNA polymerase (Promega). Amplification was done by an initial incubation at 94 °C for 2 min, followed by five cycles of incubation at 94 °C for 30 s, 52 °C for 30 s and 72 °C for 2 min 10 s, followed by 25 cycles at 93 °C for 20 s, 59 °C for 20 s and 72 °C for 2 min. The RT-PCR products were analysed by agarose gel electrophoresis and staining with ethidium bromide.
|
RESULTS |
|---|
|
TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
|---|
). Therefore, it was considered likely that expression of these sequences might convey some advantage to the virus vector. As HCPro and NIb of potyviruses are translated as a polyprotein that is cleaved into 10 proteins (Urcuqui-Inchima et al., 2001), expression of these sequences from other viruses requires prior terminal modification of the coding sequences. Therefore, ORFs with initiator and terminator codons were constructed from the HCPro and NIb coding sequences of PVY and were inserted into the multiple cloning site of a PVX vector, to generate PVX-HCPro and PVX-NIb (Fig. 1). In addition, to examine the effect of expression of one of these genes from a different virus vector system, the PVY NIb ORF was inserted into RNA2 of a TRV vector, to generate TRV-NIb (Fig. 1). These hybrid viruses were first examined for their stability during infection over a 3 week (N. benthamiana) or 4 week (tobacco) period; the different time periods for sampling the two hosts reflect the different rates of systemic infection by the recombinant viruses. Subsequently, the TRV-NIb hybrid virus was examined for its ability to infect PVY NIb-transgenic tobacco plants resistant to infection by PVY. The wt TRV RNA2 of strain PpK20 (Hernández et al., 1995) was chosen rather than the empty TRV RNA2 vector, since wt TRV RNA2 is similar in size to TRV-NIb (Fig. 1). This is because the TRV RNA2 vector has a deletion of the complete 2c and most of the 2b genes, which are not required for replication or movement, as well as an insertion of a duplicated CP subgenomic promoter from the related tobravirus Pea early browning virus (MacFarlane & Popovich, 2000).
|
) and in N. benthamiana (Fig. 2b). However, at all three time points p.i., the levels of accumulation of the hybrid viruses were lower than for the parental virus (the empty PVX vector), regardless of the host species. Furthermore, differences in the size of the genomic RNA were apparent in some samples, and the number of samples with such shorter RNAs increased with time p.i. (Fig. 2). This was true in both species. Although the appearance of smaller-sized RNAs in tobacco occurred at 2 weeks p.i. in one out of four plants infected by either PVX-NIb or PVX-HCPro, in subsequent time samples, the smaller-sized, full-length RNA appeared in more of the samples infected with PVX expressing the HCPro gene than in those infected with PVX expressing the NIb gene (Figs 2a). That is, in tobacco at 3 weeks p.i., none of the four PVX-NIb samples contained the shorter, wt-sized (6.5 kb) RNA, while at 4 weeks p.i., two of the four samples contained the shorter genomic RNA. By contrast, at 3 weeks p.i., two of the four PVX-HCPro samples contained a mixture of both RNAs and one contained only the shorter RNA, while at 4 weeks p.i., all four samples contained only the shorter RNA. These data suggest that the hybrid viruses are not very stable during propagation in tobacco and viruses with smaller or no inserts are preferentially selected. However, the situation in N. benthamiana was less clear. In this species, although smaller-sized RNAs appeared, which were different in size from the expected subgenomic RNAs of the hybrid viruses (3.8, 3.7, 2.6 and 2.5 kb), an RNA larger in size (8.0 or 8.1 kb) than the empty PVX vector (6.5 kb) was still present in all but one sample as seen in Fig. 2(b), suggesting that selection of smaller-sized RNA species may not be as strong or rapid in this host. However, this conclusion was not supported by subsequent analysis (see below).
|
In tobacco, it was apparent that at 2 weeks p.i., very little of the RNA population in the plants infected with PVX-NIb had undergone deletion of the 1581 bp insert (Fig. 3a, lanes 9–12). At 3 weeks p.i, while there was some increase in the number and abundance of minor PCR products, all of the samples from plants infected by PVX-NIb yielded a PCR product corresponding to the full-length insert plus flanking sequences (1882 bp). However, by 4 weeks p.i., only two of the four samples still retained the band reflecting the full-length insert, while the other two samples contained inserts of smaller size, although still larger than the 301 bp product generated from the empty vector (Fig. 3a, lanes 9–12). By contrast, as in the Northern blot (Fig. 2a
), only one of the four samples taken at 2 weeks p.i. from plants infected with PVX-HCPro consisted mostly of virus with the 1455 bp HCPro insert deleted, although this sample also still contained virus with little or no deletion present in a fraction of the RNA population (Fig. 3a, lane 15). By 3 weeks p.i., only one of the four samples contained predominantly virus with the 1455 bp insert plus 295 bp flanking sequences expected of the PVX-HCPro inoculated (Fig. 3a, lane 13), while at 4 weeks p.i., none of the samples contained only PVX-HCPro with the full-length insert plus flanking sequences (Fig. 3a).
|
, lanes 9–16). However, by 3 weeks p.i., plants infected with either hybrid virus showed the presence of multiple deleted forms (Fig. 3b, lanes 9–16). In an independent experiment (Supplementary Fig. S1, available in JGV Online), both PVX-NIb and PVX-HCPro also showed the presence of some deleted forms at 1 and 2 weeks p.i, with no full-length form detectable at 3 weeks p.i. Here, in contrast to the situation in tobacco, PVX-HCPro did not appear to be either more or less stable than PVX-NIb (Figs 2a vs 2b and 3a vs 3b). The absence of any PCR product in one of the PVX-NIb samples taken at 2 weeks p.i. (Fig. 3b, lane 11) is consistent with the smaller-sized genomic RNA seen in this sample on a Northern blot (Fig. 2b, lane 8, asterisk), and may represent a sample in which one of the PCR priming sites was lost. However, it also appears that a population of molecules with some inserts remaining still existed in this plant, as the sample taken 1 week later from a different leaf of the same plant contained PVX-NIb with several intermediate-sized inserts (Fig. 3b, lane 11) as well as the larger-sized genomic RNA species (Fig. 2b, lane 8). Thus, the data showed that while the levels of virus accumulation in plants infected by PVX-HCPro and PVX-NIb were lower than for the empty PVX vector, the inserts were lost to varying extents depending on the host and, in the case of tobacco, also depending on the nature of the insert sequence. Moreover, the extent to which sequences were lost and the degree of co-existence of viruses with larger inserts and no insert varied in different plants, as well as in different leaves of the same plant and, to some degree, in different experiments.
Accumulation and stability of TRV-NIb
Infection of tobacco with TRV-NIb vs TRV and assessment of the accumulation of TRV RNA2 showed that all of the plants were infected and that the level of accumulation of TRV-NIb at 2 weeks p.i. was not substantially different from that of wt TRV RNA2 (Fig. 4a). There was considerable variation in the level of accumulation of both RNA2s in different plants and in samples taken at different times (Fig. 4a). However, the presence of smaller, abundant PCR products in the samples taken from plants infected by TRV-NIb suggested that the full-length insert was not maintained stably in tobacco. This was verified by an RT-PCR analysis of the same samples (Fig. 4b). Here again, some samples did not yield detectable PCR products, suggesting that one of the primer sites may have been lost due to the deletion. While a mixed population of viruses with full-length (2046 bp) and smaller inserts were present at 2 weeks p.i., the majority of the virus population appeared to have lost most or all of their inserts at 3 and 4 weeks p.i. (Figs 4a, b).
|
). This was confirmed by the RT-PCR analysis (Fig. 5b). It does not appear from the RT-PCR analyses that, in general, inserts of larger size were retained longer in N. benthamiana than in tobacco (Fig. 5b vs Fig. 4b).
|
) and either susceptible or resistant to infection by PVY. No systemic infection was observed in the NIb-transgenic tobacco resistant to PVY (not shown). Therefore, inoculated leaves were sampled for accumulation of TRV at 4 days p.i. The wt TRV accumulated in non-transgenic tobacco and NIb-transgenic tobacco susceptible or resistant to PVY (Fig. 6a). There was some variation in the levels of accumulation between different plants of the same genotype treated with the same inoculum, but the intact genomic RNA2 was the major form of the TRV RNA2 present (Fig. 6a). Similarly, TRV-NIb inoculated to non-transgenic tobacco showed equivalent levels of accumulation of the genomic TRV RNA2, also with some variation in different plants of the same genotype (Fig. 6a). However, TRV-NIb inoculated to NIb-transgenic tobacco susceptible or resistant to PVY showed a different pattern of accumulation. In the PVY-susceptible NIb-transgenic plants, TRV-NIb accumulated to either similar or much lower levels than in the non-transgenic plants, with only the former plants retaining the full-length TRV-NIb as the major species (Fig. 6a). By contrast, in the PVY-resistant NIb-transgenic plants, TRV RNA2 with the full-length NIb insert did not accumulate and only deletion variants of the TRV-NIb RNA2 accumulated. In several plants, no TRV-NIb RNA2 was detectable (Fig. 6a and data not shown). These conclusions were confirmed by RT-PCR analysis of the same samples (Fig. 6b and data not shown). Therefore, as expected, the loss of the NIb sequence was necessary for accumulation of TRV RNA2 in the PVY-resistant NIb-transgenic plants, and the resistance mechanism increased the rate of selection of the deleted variants, which were then detectable in a few days rather than weeks.
|
|
DISCUSSION |
|---|
|
TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
|---|
; Canto et al., 2004; Li et al., 1999; Pruss et al., 1997; Scholthof et al., 1995; Thomas et al., 2003). In many cases, the hybrid viruses induced necrosis in some hosts and, in one case, the hybrid virus induced an avirulent response limiting infection by that virus (Li et al., 1999). However, these viruses were not assessed for their competitiveness with their parental viruses. In one case where the hybrid virus [Tobacco mosaic virus (TMV) with the capsid protein of Alfalfa mosaic virus] was not more pathogenic than either parental virus, infection of a new host species (for TMV) was observed by the hybrid virus, although it was also shown not to be as fit as the parental virus (TMV) during infection of a common host (Spitsin et al., 1999).
Although expression of the PVY HCPro and NIb has been reported to enhance the pathogenicity of PVX (Brigneti et al., 1998), we did not observe such effects consistently in this series of experiments. This may have been because our analyses have shown that the HCPro and NIb sequences of PVY were not maintained stably in PVX in either of two hosts, N. tabacum or N. benthamiana. While some differences were noted in the rate of selection of viruses with most or all of the sequences deleted in one host vs another or depending on the transgene, overall, the effect was the same. That is, the hybrid viruses did not accumulate to as high a level as the empty virus vector and, several weeks p.i., the hybrid viruses had lost most or all of the inserted sequences (Figs 2, 3). A similar result over a similar time frame was observed with PVX expressing a mutant form of the plum pox virus (PPV) HCPro in N. benthamiana (Barajas et al., 2006).
In the case of TRV RNA2 expressing the NIb sequence, the hybrid virus accumulated to similar levels to the wt TRV RNA2 and some of the hybrid virus was still present in N. benthamiana plants at the last time point tested. Nevertheless, virus with little or no insert accumulated preferentially in both hosts tested (Figs 4, 5). Moreover, in transgenic plants resistant to PVY, mediated by RNA silencing of the NIb transgene, TRV-NIb was strongly selected against, with the virus either losing the inserted sequence or not accumulating, as it was now targeted by the resistance against PVY (NIb). Thus, after they are initially formed, it is difficult to see how hybrid viruses could accumulate to detectable levels during competition for further infection with their parental viruses.
The inability of PVX expressing a viral sequence to retain the insert and accumulate in some transgenic plant lines expressing the same viral sequence was also observed in two recent studies (Barajas et al., 2006; Roy et al., 2006). In both of these studies, only PVX with deleted forms of the insert was able to accumulate. In one case, the nature of the partial deletions was examined and found to relate to the regions of the inserts targeted by the small interfering RNAs generated from the transgene (Barajas et al., 2006). Although we did not see such deleted TRV variants arise in the case of the transgenic plants resistant to PVY (Fig. 6), the extent to which different hybrid viruses are able to survive sufficiently to generate such deleted variants seems to depend on the strength of the resistance (Barajas et al., 2006).
There are several conclusions that can be drawn from this and previous studies. (i) Some, if not most, hybrid viruses formed by recombination between a viral transgene and a non-target virus are unlikely to be as fit as the parental, non-target virus, even if they accumulate initially to similar levels. This is supported by several studies showing the instability of some viral vectors expressing non-viral sequences (Chapman et al., 1992; Dolja et al., 1993; Guo et al., 1998; Rabindran & Dawson, 2001). (ii) The size of the insert may be an important factor in the stability of the hybrid virus, since hybrid viruses with shorter remaining inserts persisted longer than the hybrid virus with the full-length insert, although these too were not competitive with the wt virus. This conclusion is similar to those of others using viral vectors to express non-viral sequences (Chapman et al., 1992; Dawson et al., 1989; Dolja et al., 1993; Guo et al., 1998; Rabindran & Dawson, 2001). (iii) While a hybrid virus could in principle be generated by recombination between two co-infecting viruses, this has not been seen to date for unrelated viruses, probably since these novel, hybrid viruses would be present at very low levels and cannot compete efficiently with the parental viruses. (iv) The concern about generation of novel, hybrid viruses by recombination between a viral transgene and a non-target virus can be mitigated to some extent by the observation that the hybrid virus itself is a target of the resistance response. This also was seen in two recent studies in which such hybrid viruses were used to characterize the resistance (Barajas et al., 2006; Roy et al., 2006). Moreover, if the non-target parental virus inhibited RNA silencing-mediated resistance in the infected tissues (which may need to be a pre-requisite for enough transgene mRNA to accumulate for recombination) so that the hybrid virus could accumulate, the latter might then be non-competitive with the non-target parental virus. If somehow the hybrid virus did accumulate and escape to adjacent plants, then it would be sensitive to the RNA silencing-mediated resistance in those plants or, if those plants were pre-infected with the non-target parental virus, the hybrid virus would be subject to cross-protection by that parental virus. Thus, although situations may exist that will favour the hybrid virus, neither of the above circumstances, would facilitate such a hybrid virus becoming established. Data generated using other viral vector/viral inserts will determine how general these conclusions are and whether or not there are exceptions.
|
ACKNOWLEDGEMENTS |
|---|
|
REFERENCES |
|---|
|
TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
|---|
Adair, T. L. & Kearney, C. M. (2000). Recombination between a 3-kilobase tobacco mosaic virus transgene and a homologous viral construct in the restoration of viral and nonviral genes. Arch Virol 145, 1867–1883.[CrossRef][Medline]
Audy, P., Palukaitis, P., Slack, S. A. & Zaitlin, M. (1994). Replicase-mediated resistance to potato virus Y in transgenic tobacco plants. Mol Plant Microbe Interact 7, 15–22.[Medline]
Barajas, D., Tenllado, F. & Diaz-Ruiz, J. R. (2006). Characterization of the recombinant forms arising from a Potato virus X chimeric virus infection under RNA silencing pressure. Mol Plant Microbe Interact 19, 904–913.[Medline]
Baulcombe, D. C., Chapman, S. & Santa Cruz, S. (1995). Jellyfish green fluorescent protein as a reporter for virus infection. Plant J 7, 1045–1053.[CrossRef][Medline]
Bejarano, E. R., Khashoggi, A., Witty, M. & Lichtenstein, C. (1996). Integration of multiple repeats of geminiviral DNA into the nuclear genome of tobacco during evolution. Proc Natl Acad Sci U S A 93, 759–764.
Borja, M., Rubio, T., Scholthof, H. B. & Jackson, A. O. (1999). Restoration of wild-type virus by double recombination of tombusvirus mutants with host transgene. Mol Plant Microbe Interact 12, 153–162.[Medline]
Bousalem, M., Douzery, E. J. & Fargette, D. (2000). High genetic diversity, distant phylogenetic relationships and intraspecies recombination events among natural populations of Yam mosaic virus: a contribution to understanding potyvirus evolution. J Gen Virol 81, 243–255.
Brigneti, G., Voinnet, O., Li, W.-X., Ji, L.-H., Ding, S.-W. & Baulcombe, D. C. (1998). Viral pathogenicity determinants are suppressors of transgene silencing in Nicotiana benthamiana. EMBO J 17, 6739–6746.[CrossRef][Medline]
Burgyán, J., Hornyik, C., Szittya, G., Silhavy, D. & Bisztray, G. (2000). The ORF1 products of tombusviruses play a crucial role in lethal necrosis in virus-infected plants. J Virol 74, 10873–10881.
Canto, T. & Palukaitis, P. (1998). Transgenically expressed cucumber mosaic virus RNA 1 simultaneously complements replication of cucumber mosaic virus RNAs 2 and 3 and confers resistance to systemic infection. Virology 250, 325–336.[CrossRef][Medline]
Canto, T. & Palukaitis, P. (2001). A cucumber mosaic virus (CMV) RNA 1 transgene mediates suppression of the homologous viral RNA 1 constitutively and prevents CMV entry into the phloem. J Virol 75, 9114–9120.
Canto, T. & Palukaitis, P. (2002). Novel N gene-associated, temperature-independent resistance to the movement of Tobacco mosaic virus vectors neutralized by a Cucumber mosaic virus RNA1 transgene. J Virol 76, 12908–12916.
Canto, T., MacFarlane, S. A. & Palukaitis, P. (2004). ORF6 of Tobacco mosaic virus is a pathogenicity determinant in Nicotiana benthamiana. J Gen Virol 85, 3123–3133.
Cervera, M. T., Riechmann, J. L., Martin, M. T. & Garcia, J. A. (1993). 3'-Terminal sequence of the plum pox virus PS and o6 isolates: evidence for RNA recombination within the potyvirus group. J Gen Virol 74, 329–334.
Chapman, S., Kavanagh, T. & Baulcombe, D. (1992). Potato virus X as a vector for gene expression in plants. Plant J 2, 549–557.[Medline]
Chare, E. R. & Holmes, E. C. (2006). A phylogeny survey of recombination frequency in plant RNA viruses. Arch Virol 151, 933–946.[CrossRef][Medline]
Dawson, W. O., Lewandowski, D. J., Hilf, M. E., Bubrick, P., Raffo, A. J., Shaw, J. J., Grantham, G. L. & Desjardins, P. R. (1989). A tobacco mosaic virus-hybrid expresses and loses an added gene. Virology 172, 285–292.[CrossRef][Medline]
Desbiez, C. & Lecoq, H. (2004). The nucleotide sequence of Watermelon mosaic virus (WMV, Potyvirus) reveals interspecific recombination between two related potyviruses and the 5' part of the genome. Arch Virol 149, 1619–1632.[Medline]
Dolja, V. V., Herndon, K. L., Pirone, T. P. & Carrington, J. C. (1993). Spontaneous mutagenesis of a plant potyvirus genome after insertion of a foreign gene. J Virol 67, 5968–5975.
Glais, L., Tribodet, M. & Kerlan, C. (2002). Genomic variability in Potato potyvirus Y (PVY): evidence that PVYNW and PVYNTN variants are single multiple recombinants between PVYO and PVYN isolates. Arch Virol 147, 363–378.[CrossRef][Medline]
Goldbach, R., Bucher, E. & Prins, M. (2003). Resistance mechanisms to plant viruses: an overview. Virus Res 92, 207–212.[CrossRef][Medline]
Greene, A. E. & Allison, R. F. (1994). Recombination between viral RNA and transgenic plant transcripts. Science 263, 1423–1425.
Greene, A. E. & Allison, R. F. (1996). Deletions in the 3' untranslated region of Cowpea chlorotic mottle virus transgene reduce recovery of recombinant viruses in transgenic plants. Virology 225, 231–234.[CrossRef][Medline]
Guo, H. S., Lopez-Moya, J. J. & Garcia, J. A. (1998). Susceptibility to recombination rearrangement of a chimeric plum pox potyvirus genome after insertion of a foreign gene. Virus Res 57, 183–195.[CrossRef][Medline]
Harper, G., Osuji, J. O., Heslop-Harrison, J. S. & Hull, R. (1999). Integration of banana streak badnavirus into the Musa genome: molecular and cytogenetic evidence. Virology 255, 207–213.[CrossRef][Medline]
Hernández, C., Mathis, A., Brown, D. J. F. & Bol, J. F. (1995). Sequences of RNA 2 of a nematode-transmissible isolate of tobacco rattle virus. J Gen Virol 76, 2847–2851.
Jakowitsch, J., Mette, M. F., van der Winden, J., Matzke, M. A. & Matzke, A. J. M. (1999). Integrated pararetroviral sequences define a unique class of dispersed repetitive DNA in plants. Proc Natl Acad Sci U S A 96, 13241–13246.
Krause-Sakate, R., Fakhfakh, H., Peypelut, M., Pavan, M. A., Zerbini, F. M., Marrakchi, M., Candresse, T. & Le Gall, O. (2004). A naturally occurring recombinant isolate of Lettuce mosaic virus. Arch Virol 149, 191–197.[CrossRef][Medline]
Larsen, R. C., Miklas, P. N., Druffel, K. L. & Wyatt, S. D. (2005). NL-3 K strain is a stable and naturally occurring interspecific recombinant derived from Bean common mosaic necrotic virus and Bean common mosaic virus. Phytopathology 95, 1037–1042.
Li, H.-W., Lucy, A. P., Guo, H.-S., Li, W.-X., Ji, L.-H., Wong, S.-M. & Ding, S.-W. (1999). Strong host resistance targeted against a viral suppressor of the plant gene silencing defence mechanism. EMBO J 18, 2683–2691.[CrossRef][Medline]
MacFarlane, S. A. & Popovich, A.-H. (2000). Expression of foreign proteins in roots from tobravirus vectors. Virology 267, 29–35.[CrossRef][Medline]
Masuta, C., Ueda, S., Suzuki, M. & Uyeda, I. (1998). Evolution of a quadripartite hybrid virus by interspecific exchange and recombination between replicase components of two related tripartite RNA viruses. Proc Natl Acad Sci U S A 95, 10487–10492.
Mueller, A.-M., Mooney, A. L. & MacFarlane, S. A. (1997). Replication of in vitro tobravirus recombinants shows that the specificity of template recognition is determined by 5' non-coding but not 3' non-coding sequences. J Gen Virol 78, 2085–2088.[Abstract]
Palukaitis, P. (1991). Virus-mediated genetic transfer in plants. In Risk Assessment in Genetic Engineering: Environmental Release of Organisms, pp. 140–162. Edited by M. Levin & H.S. Strauss. New York: McGraw Hill.
Palukaitis, P. & Kaplan, I. B. (1997). Synergy of virus accumulation and pathology in transgenic plants expressing viral sequences. In Virus-Resistant Transgenic Plants: Potential Ecological Impact, pp. 77–84. Edited by M. Tepfer & E. Balázs. Berlin: Springer.
Prins, M. (2003). Broad virus resistance in transgenic plants. Trends Biotechnol 21, 373–375.[CrossRef][Medline]
Pruss, G., Ge, X., Shi, X. M., Carrington, J. C. & Vance, V. B. (1997). Plant viral synergism: the potyviral genome encodes a broad-range pathogenicity enhancer that transactivates replication of heterologous viruses. Plant Cell 9, 859–868.
Rabindran, S. & Dawson, W. O. (2001). Assessments of recombinants that arise from the use of a TMV-based transient expression vector. Virology 284, 182–189.[CrossRef][Medline]
Revers, F., Le Gall, O., Candresse, T., Le Romancer, M. & Dunez, J. (1996). Frequent occurrence of recombinant potyvirus isolates. J Gen Virol 77, 1953–1965.
Roy, G., Sudarshana, M. R., Ullman, D. E., Ding, S.-W., Dandekar, A. M. & Falk, B. W. (2006). Chimeric cDNA sequences from Citrus tristeza virus confer RNA silencing-mediated resistance in transgenic Nicotiana benthamiana plants. Phytopathology 96, 819–827.
Schoelz, J. E. & Wintermantel, W. M. (1993). Expansion of viral host range through complementation and recombination in transgenic plants. Plant Cell 5, 1669–1679.[Abstract]
Scholthof, H. B., Scholthof, K.-B. G. & Jackson, A. O. (1995). Identification of tomato bushy stunt virus host-specific symptom determinants by expression of individual genes from a potato virus X vector. Plant Cell 7, 1157–1172.[Abstract]
Spitsin, S., Steplewski, K., Fleysh, N., Belanger, H., Mikheeva, T., Shivprasad, S., Dawson, W., Koprowski, H. & Yusibov, V. (1999). Expression of alfalfa mosaic virus coat protein in tobacco mosaic virus (TMV) deficient in the production of its native coat protein supports long-distance movement of a chimeric TMV. Proc Natl Acad Sci U S A 96, 2549–2553.
Tan, Z., Wada, Y., Chen, J. & Ohshima, K. (2004). Inter- and intralineage recombinants are common in natural populations of Turnip mosaic virus. J Gen Virol 85, 2683–2696.
Tanne, E. & Sela, I. (2005). Occurrence of a DNA sequence of a non retro RNA virus in a host genome and its expression: evidence of recombination between viral and host RNAs. Virology 332, 614–622.[CrossRef][Medline]
Tepfer, M. (2002). Risk-assessment of virus resistant transgenic plants. Annu Rev Phytopathol 40, 467–491.[CrossRef][Medline]
Thomas, C. L., Leh, V., Lederer, C. & Maule, A. J. (2003). Turnip crinkle virus coat protein mediates suppression of RNA silencing in Nicotiana benthamiana. Virology 306, 33–41.[CrossRef][Medline]
Uhrig, J. F., Canto, T., Marshall, D. & MacFarlane, S. A. (2004). Relocalization of nuclear ALY proteins to the cytoplasm by the tomato bushy stunt virus P19 pathogenicity protein. Plant Physiol 135, 2411–2423.
Urcuqui-Inchima, S., Haenni, A.-L. & Bernardi, F. (2001). Potyvirus proteins: a wealth of functions. Virus Res 74, 157–175.[CrossRef][Medline]
Varrelmann, M., Palkovics, L. & Maiss, E. (2000). Transgenic or plant expression-vector mediated recombination of Plum pox virus. J Virol 74, 7462–7469.
White, K. A. & Morris, T. J. (1994). Recombination between defective tombusvirus RNAs generates functional hybrid genomes. Proc Natl Acad Sci U S A 91, 3642–3646.
Wintermantel, W. M. & Schoelz, J. E. (1996). Isolation of recombinant viruses between cauliflower mosaic virus and a viral gene in transgenic plants under conditions of moderate selection pressure. Virology 223, 156–164.[CrossRef][Medline]
Zhong, Y., Guo, A., Li, C., Zhuang, B., Lai, M., Wei, C., Luo, J. & Li, Y. (2005). Identification of a naturally occurring recombinant isolate of Sugarcane mosaic virus causing maize dwarf mosaic disease. Virus Genes 30, 75–83.[CrossRef][Medline]
Received 16 August 2006;
accepted 29 November 2006.
This article has been cited by other articles:
|
|
 |
|
  W. J. Cuellar, F. Tairo, J. F. Kreuze, and J. P. T. Valkonen 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 J. Gen. Virol., February 1, 2008; 89(2): 573 - 582. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| INT J SYST EVOL MICROBIOL | MICROBIOLOGY | J GEN VIROL |
| J MED MICROBIOL | ALL SGM JOURNALS | |
