Phylogenetic analysis of some large double-stranded RNA replicons from plants suggests they evolved from a defective single-stranded RNA virus

Mark J. Gibbs1, Ryuichi Koga2, Hiromitsu Moriyamab,2, Pierre Pfeiffer3 and Toshiyuki Fukuhara2

Bioinformatics, Research School of Biological Sciences, The Australian National University, GPO Box 475, Canberra 2601, Australia1
Laboratory of Molecular Cell Biology, Faculty of Agriculture, Tokyo University of Agriculture and Technology, Fuchu, Tokyo 183, Japan2
Institute of Plant Molecular Biology, 12 Rue du Général Zimmer, 67000 Strasbourg, France3

Author for correspondence: Mark Gibbs. Fax +61 2 62494437. e-mail mgibbs{at}rsbs.anu.edu.au


   Abstract
Top
Abstract
Introduction
Methods
Results and Discussion
References
 
Sequences were recently obtained from four double-stranded (ds) RNAs from different plant species. These dsRNAs are not associated with particles and as they appeared not to be horizontally transmitted, they were thought to be a kind of RNA plasmid. Here we report that the RNA-dependent RNA polymerase (RdRp) and helicase domains encoded by these dsRNAs are related to those of viruses of the alpha-like virus supergroup. Recent work on the RdRp sequences of alpha-like viruses raised doubts about their relatedness, but our analyses confirm that almost all the viruses previously assigned to the supergroup are related. Alpha-like viruses have single-stranded (ss) RNA genomes and produce particles, and they are much more diverse than the dsRNAs. This difference in diversity suggests the ssRNA alpha-like virus form is older, and we speculate that the transformation to a dsRNA form began when an ancestral ssRNA virus lost its virion protein gene. The phylogeny of the dsRNAs indicates this transformation was not recent and features of the dsRNA genome structure and translation strategy suggest it is now irreversible. Our analyses also show some dsRNAs from distantly related plants are closely related, indicating they have not strictly co-speciated with their hosts. In view of the affinities of the dsRNAs, we believe they should be classified as viruses and we suggest they be recognized as members of a new virus genus (Endornavirus) and family (Endoviridae).


   Introduction
Top
Abstract
Introduction
Methods
Results and Discussion
References
 
Complete sequences have been obtained from large double-stranded (ds) RNA species found in cultivated rice (Oryza sativa ssp. Japonica; Moriyama et al., 1995 ), wild rice (Oryza rufipogon;Moriyama et al., 1999 ) and broad bean (Vicia faba cv. 447; Pfeiffer, 1998 ) and a partial sequence has been obtained for one of two possible species found in kidney bean (Phaselous vulgaris cv. black turtle soup; Wakarchuk & Hamilton, 1990 ). Similar large dsRNAs have been found in a few other plant species, and together these dsRNAs have been called ‘endogenous dsRNAs’ (Moriyama et al., 1996 , 1999 ). They have an unusual combination of properties. They are similar to cryptoviruses (Ghabrial et al., 1995 ) in that they are efficiently transmitted through seed, no horizontal spread has been observed in the field, no potential vectors have been identified and none is associated with disease symptoms, except for one associated with sterility (Pfeiffer et al., 1993 ). However, unlike cryptoviruses, which produce particles and have a genome consisting of two dsRNAs each about 2–3 kb long, none of the endogenous dsRNAs is associated with particles and each is more than 10 kb long. Consistent with a lack of particles, none of the endogenous dsRNAs is mechanically transmissible.

Work on the dsRNA from O. sativa showed that it is not encoded by the host and that its replication is regulated to produce a constant low concentration in all tissues except pollen (Fukuhara et al., 1993 ; Moriyama et al., 1996 , 1999 ). Some of the other dsRNAs have also been found at a constant, low concentration in their hosts, suggesting that this is a group characteristic (Wakarchuk & Hamilton, 1985 ; Gabriel et al., 1987 ; Fairbanks et al., 1988 ; Valverde & Fontenot, 1990 ; Zabalgogeazcoa & Gildow, 1992 ). This regulation, the apparent non-infectious nature of the dsRNAs, and their lack of particles led Fukuhara et al. (1993) to suggest that these organisms are a kind of RNA plasmid. Studies on the V. faba dsRNA showed that it does not even spread from cell to cell except at cell division (Duc et al., 1984 ), supporting this view. Fukuhara et al. (1993) also suggested that the endogenous dsRNAs might be related to hypoviruses as these fungal viruses similarly lack particles and are transmitted vertically but not horizontally except by hyphal fusion. Hypoviruses also have large dsRNA genomes and they too were once considered to be a kind of RNA plasmid (Brown & Finnegan, 1989 ; Shapira et al., 1991 ).

Here we report an analysis of the sequences of the endogenous dsRNAs that clarifies their relationships, provides new information on their evolution and supports a clear argument for their classification. We also provide new evidence that supports grouping a wide set of viruses, including the dsRNAs, into the previously debated alpha-like virus supergroup.


   Methods
Top
Abstract
Introduction
Methods
Results and Discussion
References
 
{blacksquare}
The non-redundant coding and nucleotide sequence databases were searched using the programs BLASTP, TBLASTN and PSI-BLAST (Altschul et al., 1997 ) and Expect values (E values) were calculated by these programs for pairs of aligned sequences. The evolutionary relatedness (homology) of sequences was judged firstly using these E values. E values are estimates of the probability of finding the same degree of similarity by chance given the composition of the aligned sequences and the database. E values less than 1x10-2 usually indicate homology and those less than 1x10-6 almost always indicate homology (Altschul et al., 1997 ; Bork & Gibson, 1996 ). The similarity of pairs of sequences was also measured in percent identity plots made using the program PLOTSIMILARITY (Devereux et al., 1984 ) and using the program ALIGN (Dayhoff et al., 1983 ), which was used to make a second test of relatedness. ALIGN performs a Monte Carlo procedure to calculate a Z-score which is largely unbiased by sequence composition and length. Usually a Z-score of 3 or more would be taken to be significant, but because of the constraints on real proteins, alignments of their sequences produce biased scores centred on 3 rather than 0 (Barton & Sternberg, 1987 ). Thus, it is generally accepted that Z-scores of 5 to 6 indicate relatedness (Barton, 1996 ) when produced by this method. We did 100 alignments from randomized sequences for each pairwise comparison made with the program ALIGN and used the MDM78 distance matrix (Dayhoff et al., 1978 ) and a gap penalty of 4 in the calculations.

Multiple alignments of amino acid and nucleotide sequences were made with the program CLUSTALW (Thompson et al., 1994 ). Maximum likelihood trees were found from the amino acid multiple alignments by quartet puzzling using the program PUZZLE version 4 (Strimmer & von Haeseler, 1996 ) after positions including gaps had been excluded. Likelihoods were calculated using the BLOSUM 62 substitution matrix (Henikoff & Henikof, 1992 ) and a gamma distribution of rates of change for variable sites with a shape parameter estimated from the data using a neighbour-joining tree. Maximum likelihood and most parsimonious trees were also found from the aligned nucleotide sequences by heuristic searching with the program PAUP version 4d64 (written by David L. Swofford) after positions including gaps and third codon positions had been excluded. Bootstrap values were calculated from neighbour-joining trees inferred from 1000 bootstrap samples.


   Results and Discussion
Top
Abstract
Introduction
Methods
Results and Discussion
References
 
Intra-group homology and phylogeny
Each of the three completely sequenced dsRNAs encodes a single long open reading frame which we will call the long protein (LP) gene. Comparisons of the LP amino acid sequences from the O. sativa and V. faba dsRNAs showed that the likely helicase and RNA-dependent RNA polymerase (RdRp) domains correspond to two stretches of higher identity (Fig. 1, bottom curve). These two regions were also identified in database searches (Fig. 1). An E value of 3x10-22 was obtained for an alignment 670 residues long between the regions that include the helicase motifs from the V. faba and O. sativa dsRNA LPs. An E value of 2x10-75 was obtained for an alignment 370 residues long between the regions that include the RdRp motifs from these same two LP sequences. These two database search results independently indicated that the Oryza and V. faba dsRNAs share a common ancestor.



View larger version (16K):
[in this window]
[in a new window]
 
Fig. 1. Percent identity plots found using a window 100 amino acid residues long for an alignment of the long proteins (LPs) of the two Oryza dsRNAs (top curve) and for an alignment of the LPs of the O. sativa and V. faba dsRNAs (lower curve). The upper curve is scaled to the upper x-axis and the lower curve is scaled to the lower x-axis. The conserved GKT/S motif (marked with open triangles) is close to the beginning of the helicase domain, and the conserved GDD motif (marked with open circles) is close to the end of the RdRp domain. Black filled blocks mark similar regions between the V. faba and O. sativa dsRNAs identified by a BLASTP search. E values obtained from the alignments of these regions are shown. Note that the LPs of the Oryza dsRNAs are about 4600 amino acid residues long and the LP of the V. faba dsRNA is 5825 residues long.

 
Database searches with the TBLASTN program using LP sequences from the Oryza dsRNAs identified similarities with the partial sequence of a dsRNA from P. vulgaris. An alignment of the P. vulgaris polypeptide with the LP from the O. sativa dsRNA yielded an E value of 1x10-10. By contrast, database searches with the LP sequence from the V. faba dsRNA did not detect similarities with the P. vulgaris dsRNA sequence. We confirmed this difference in affinities using the Monte Carlo procedure. Z-scores of 12·5 and 11·2 were obtained for comparisons between amino acid sequences encoded by the P. vulgaris and Oryza dsRNAs, but a Z-score of only 4·2 was obtained for the comparison of the P. vulgaris dsRNA sequence and the equivalent sequence from the V. faba dsRNA. A maximum-likelihood tree inferred from these sequences concurred with this result (Fig. 2, top). As our sequence comparisons showed that the V. faba and Oryza dsRNAs have a common ancestor, and that the P. vulgaris and Oryza dsRNAs are similarly related, we conclude that the P. vulgaris and V. faba dsRNAs are also related. A multiple alignment supported this conclusion as it showed that 22% of the amino acid residues were strictly or strongly conserved in the equivalent P. vulgaris, Oryza and V. faba dsRNA LP sequences. Our conclusion is also supported by the work of Wakarchuk & Hamilton (1985) , who showed that the P. vulgaris dsRNA shares several properties with the V. faba and Oryza dsRNAs. Together these data suggest the genome of the P. vulgaris dsRNA is similar to those of the fully sequenced dsRNAs and on this basis we suggest the P. vulgaris dsRNA should be placed in the same taxonomic family.



View larger version (17K):
[in this window]
[in a new window]
 
Fig. 2. A maximum likelihood tree of the RNA-dependent RNA polymerases of the alpha-like viruses and the endogenous dsRNAs. The tree was found from aligned amino acid sequences. Maximum likelihood and most parsimonious trees found from aligned nucleotide sequences matched the tree shown. A maximum likelihood tree constructed from the helicase domain sequences also matched the tree shown, except that the branches marked with asterisks were not found. The three clusters that are identified in the tree correspond to the three clusters identified in the text and in Table 1. The root of the tree was found using the equivalent RdRp amino acid sequences of the following carmo-like viruses: Barley yellow dwarf luteovirus PAV, Carnation mottle carmovirus, Carrot mottle mimic umbravirus, Maize chlorotic mottle machlomovirus, Tobacco necrosis necrovirus and Tomato bushy stunt tombusvirus. The actual estimate for the length of the branch leading to the carmo-like outgroup (dotted line) is double that shown. The scale relates branch lengths to the number of substitutions per site. Taxa: ACLSV, Apple chlorotic leaf spot trichovirus; AMV, Alfalfa mosaic alfamovirus; ASGV, Apple stem grooving capillovirus; BMV, Brome mosaic bromovirus; BNYVV, Beet necrotic yellow vein benyvirus; BSMV, Barley stripe mosaic hordeivirus; BYSV, Beet yellow stunt closterovirus; BYV, Beet yellows closterovirus; BVQ, Beet Q pomovirus; CMV, Cucumber mosaic cucumovirus; CTV, Citrus tristeza closterovirus; GVA, Grapevine vitivirus A; HaSV, Helicoverpa armigera stunt tetravirus; HEV, Hepatitis E virus; LCV, Little cherry closterovirus; LIYV, Lettuce infectious yellows crinivirus; OBDV, Oat blue dwarf marafivirus; ORV, Oryza rufipogon endornavirus; OSV, Oryza sativa endornavirus; PCV, Peanut clump pecluvirus; PVM, Potato M carlavirus; PVuV, Phaseolus vulgaris endornavirus; PVX, Potato X potexvirus; RBDV, Raspberry bushy dwarf idaeovirus; RuBV, Rubella rubivirus; SBWMV, Soil-borne wheat mosaic furovirus; SINV, Sindbis alphavirus; TMV, Tobacco mosaic tobamovirus; TRV, Tobacco rattle tobravirus; TSV, Tobacco streak ilarvirus; TYMV, Turnip yellow mosaic tymovirus; VFV, Vicia faba endornavirus. The inset (top) shows a mid-point rooted tree for the dsRNAs inferred from the partial sequence from the P. vulgaris dsRNA and the equivalent sequences from the other dsRNAs. Terminal nodes for the dsRNAs have been labelled with the acronyms we have suggested.

 

View this table:
[in this window]
[in a new window]
 
Table 1. Z-scores from pairwise comparisons of RdRp sequences

 
Experiments suggest that the dsRNAs are only vertically transmitted through seed, either through ovules (Duc et al., 1984 ) or through both ovules and pollen (Moriyama et al., 1996 ). Attempts have been made to transmit them mechanically, by grafting, using aphids and using dodder, but all have failed (Turpen et al., 1988 ; Valverde et al., 1990 ; Zabalgogeazcoa & Gildow, 1992 ). An implication of this likely limit on spread is that the dsRNAs should co-speciate with their hosts. Endogenous dsRNAs from closely related plants should be more closely related to each other than to endogenous dsRNAs from distantly related plants. Hence, the relationships between the Oryza, P. vulgaris and V. faba dsRNAs were unexpected: Phaseolus and Vicia belong to the legume subfamily Papilionoideae and are only distant relatives of the Oryza species.

RNA-dependent RNA polymerase affinities
Surprisingly BLASTP database searches revealed similarities between the RdRp-like regions in the LP sequences and the RdRp sequences of several single-stranded (ss) RNA viruses. E values as low as 7x10-10 were obtained for alignments about 300 residues long with RdRp sequences from closteroviruses. Alignments with RdRp sequences from Hepatitis E virus (HEV) and Alfalfa mosaic alfamovirus (AMV) yielded the next lowest E values, starting at 1x10-4 and 2x10-4 respectively.

It has been suggested that the closteroviruses and alfamoviruses belong to the ‘alpha-like supergroup’ of positive-sense ssRNA viruses. This supergroup is said to also include viruses from the following genera: Alphavirus, Benyvirus, Bromovirus, Crinivirus, Cucumovirus, Capillovirus, Carlavirus, Furovirus, Hordeivirus, Idaeovirus, Marafivirus, Pecluvirus, Pomovirus, Potexvirus, Rubivirus, Tetravirus, Tobamovirus, Tobravirus, Trichovirus, Tymovirus and Vitivirus. The supergroup was defined on the basis of several features (Goldbach & de Haan, 1994 ; Koonin & Dolja, 1993 ), but only some of these are shared by all the proposed members: (i) a positive-strand RNA genome with a 5' cap, (ii) production of a subgenomic RNA encoding a virion protein, (iii) homologous RdRp and helicase amino acid sequences. Viruses that are not assigned to the supergroup share the first two of these features and hence, the supergroup is probably best defined only by the proposed homology of the helicase and RdRp sequences.

Zanotto et al. (1996) tested the support from RdRp sequence alignments for several proposed supergroups of RNA viruses and found only weak support for the alpha-like supergroup. We repeated part of this work by compiling a dataset that included at least one RdRp sequence from each genus in the supergroup for which sequence is available, and then by obtaining Z-scores by the Monte Carlo procedure. The RdRp-like regions of the endogenous dsRNAs were included in the analysis to test their relationships, as was the RdRp sequence of HEV. Sequences from carmo-like and picorna-like viruses were also included as they were likely to be outliers. As shown in Table 1, Z-scores of more than 6 were obtained for many of the comparisons. These scores proved the relatedness of almost all the RdRp sequences from viruses previously assigned to the alpha-like supergroup and showed that the RdRp sequences of the dsRNAs and HEV are related to alpha-like virus sequences. Only the affinities of the RdRp sequence of Beet necrotic yellow vein benyvirus (BNYVV), which was previously assigned to the alpha-like supergroup, were doubtful. The highest Z-score obtained for a comparison between the BNYVV sequence and an alpha-like virus sequence was only 5·8. Low Z-scores were obtained from comparisons between alpha-like virus RdRp sequences and those of carmo-like and picorna-like viruses, supporting the view that the alpha-like virus RdRp sequences are a discrete natural grouping.

Phylogenetic trees were constructed for the RdRp amino acid sequences (Fig. 2). Strong support was found in the trees for two major clusters that included RdRp sequences from the following viruses: (1) alfamoviruses, bromoviruses, closteroviruses, criniviruses, cucumoviruses, furoviruses, hordeiviruses, idaeoviruses, ilarviruses, pecluviruses, pomoviruses, tobamoviruses and tobraviruses and (2) capilloviruses, carlaviruses, marafiviruses, potexviruses, trichoviruses, tymoviruses and vitiviruses. There was also strong support for grouping the sequences from the dsRNAs, which we have designated cluster 3 in the tree and Table 1. The sequences of BNYVV, Helicoverpa armigera stunt tetravirus (HaSV), HEV, Rubella rubivirus (RuBV) and Sindbis alphavirus (SINV) were grouped in the maximum likelihood tree, but there was only weak support for this cluster and it was not found in trees inferred by other methods. Clusters 1, 2 and 3 (defined above) were supported by the Monte Carlo procedure results (Table 1), but these results did not support the grouping of BNYVV, HaSV, HEV, RuBV and SINV. The Monte Carlo results suggested that the RdRps of the dsRNAs are most closely related to those of cluster 1 and this possibility was supported by the database searches, but the phylogenetic analysis did not resolve the relationships between the clusters. The differences between our Monte Carlo randomization results and those of Zanotto et al. (1996) probably result from comparing slightly different regions of the RdRp sequences and using different parameters for the alignments. We used a region of sequence identified by the program BLASTP as high-scoring. This region began about 170 residues on the N-terminal side of the GDD motif and ended about 60 residues on its C-terminal side. Zanotto et al. (1996) tested various regions of sequence identified in earlier publications.

Zanotto et al. (1996) not only challenged the relatedness of the alpha-like viruses but also that they are monophyletic. By contrast, our phylogenetic analysis and Monte Carlo randomization results support the monophyly of the RdRp sequences from the alpha-like supergoup if this grouping includes the sequences from the dsRNAs and HEV (Fig. 2 and Table 1). One database search result challenged this conclusion: an alignment between an alpha-like RdRp sequence and that of the carmo-like virus Maize chlorotic mottle machlomovirus yielded an E value that could be significant (8x10-5), but we could not discern a clear phylogenetic signal that could link the carmo-like and alpha-like viruses (Table 1), and the carmo-like RdRp sequences are clearly outliers to the supergroup in the tree (Fig. 2). It is important to note that the actual estimates for the lengths of the branches leading to the carmo-like outlier sequences are double those shown (Fig. 2). Plainly our conclusions are contrary to some of those of Zanotto et al. (1996) , but we have only been concerned with the alpha-like supergroup. The fact that none of our database searches and analyses supported links to sequences other than those from the mentioned viruses and dsRNAs supports the theory of Zanotto et al. (1996) that there is no phylogenetic signal that could be used to group all extant RdRp sequences.

Helicase sequence affinities
A profile search using the PSI-BLAST and the helicase-like sequences from the V. faba and O. sativa dsRNAs detected similarities with helicase sequences from viruses in every genus in the alpha-like supergroup for which sequence is available. Comparisons with tobamovirus sequences produced the highest E values, 2x10-6, in the first iteration. Reciprocal searches made with the helicase sequence of Tobacco mosaic tobamovirus identified the V. faba dsRNA helicase in the first iteration with an E value of 5x10-8. Z-scores generated from a helicase sequence dataset compiled from alpha-like virus sequences were lower than those from the RdRp sequence dataset. A comparison of the V. faba dsRNA helicase with the helicase domain of Raspberry bushy dwarf idaeovirus yielded a Z-score of 7·1, as did a comparison with the helicase domain of Beet soil-borne furovirus. Tobamoviruses, idaeoviruses and furoviruses are placed in cluster 1 in the alpha-like virus RdRp tree (Fig. 2) and hence, these results support the notion that the dsRNAs may be most closely related to this subset within the supergroup.

The dsRNAs probably evolved from a defective alpha-like virus
Our results show that the endogenous dsRNAs share a common ancestor with alpha-like viruses. The RdRps and helicases encoded by the dsRNAs are probably functionally similar to their alpha-like virus counterparts, but there are differences between the modes of replication of these organisms. Alpha-like virus genomic RNA is present in host cells primarily as messenger-sense ssRNA (positive-strand RNA), but no full-length positive-strand RNA from an endogenous dsRNA has been unequivocally identified (Pfeiffer et al., 1993 ; Fukuhara et al., 1995 ) and work on the V. faba and P. vulgaris dsRNAs suggests these organisms do not produce full-length ssRNAs (Pfeiffer et al., 1993 ; Wakarchuk & Hamilton, 1985 ; Lefebvre et al., 1990 ). Possibly more significant is the fact that each of the three fully sequenced dsRNAs includes a break (discontinuity or nick) on the coding strand but not the negative strand (Pfeiffer et al., 1993 ; Fukuhara et al., 1995 ; Moriyama et al., 1999 ). The conservation of the break and the fact that the LP gene continues in-frame through the break in all three molecules, suggests the break is somehow causally linked to the dsRNA form.

The relationship between the endogenous dsRNAs and the ssRNA alpha-like viruses suggests that either the dsRNAs evolved from an ancestral ssRNA virus or vice versa. The phylogenetic trees found with the RdRp and helicase sequences support the first of these options, as they show the alpha-like viruses to be far more diverse than the dsRNAs (Fig. 2). If this is true, as we believe, then as all alpha-like viruses have ssRNA genomes and encode their own virion proteins and produce particles, it is likely that an ancestor of the dsRNAs also had these features. Two alternative explanations for the difference in diversity should be considered but may be dismissed. First, the difference could be because we do not have a fair measure of the diversity of the dsRNAs, but this is unlikely in view of the similarities between all the known endogenous dsRNAs and the evolutionary distance between the Oryza and V. faba dsRNAs. Second, the dsRNAs could be evolving far slower than the alpha-like viruses, but this is unlikely as it would mean that the dsRNAs were subjected to much stronger selection pressures than the alpha-like viruses and that this difference in selection has been maintained over long periods.

We found no sequence similarities with known virion proteins, suggesting the dsRNAs do not encode a virion protein, which correlates with the fact that the dsRNAs are not associated with particles. This unusual characteristic could be related to the evolution of the dsRNA form, as the relative accumulation of the positive-sense and complementary-sense (negative) RNA strands is known to be affected when positive-strand RNA plant viruses lose the function of their virion protein genes (Nassuth & Bol, 1983 ; French & Ahlquist, 1988 ; van der Kuyl et al., 1991 ). The balance of positive to negative RNA produced by an ancestor of the dsRNAs may have similarly altered when it lost its virion protein gene. The transformation to a dsRNA form probably became irreversible when the dsRNAs evolved the break in the positive strand.

Taxonomy
The endogenous dsRNAs could be classified as sub-viral agents because they lack particles, but the distinction between viruses and sub-viral agents is not entirely clear. The bacteriophage P4 and dependoviruses are classified as both sub-viral agents and viruses, and umbraviruses and hypoviruses are classified as viruses but not sub-viral agents, although they do not produce particles. One possible explanation for this confusion is that the dependoviruses, hypoviruses and umbraviruses, and the bacteriophage P4, are all related to conventional viruses (Mayo et al., 1995 ; Murant et al., 1995 ; Hillman et al., 1995 ). We know of no other sub-viral agents with this property and we view this situation as precedence for recognizing evolutionary relationships at this level of classification. On this basis we propose that the endogenous dsRNAs should be recognized as viruses because of their affinities to alpha-like viruses. As it is clear that the dsRNAs form a distinct group, we propose that they be nominated as members of a new virus genus and we suggest the name Endornavirus (endo, from Greek: within, and RNA) for this genus. The dsRNAs have no homology to recognized dsRNA viruses and because they are very different from other alpha-like viruses, we propose that they be assigned to a separate family, for which we suggest the name Endoviridae. To achieve continuity with the literature we also suggest that endornavirus species be named after the plant species in which they were first found. Thus, the dsRNAs for which we have sequence could be named Phaseolus vulgaris endornavirus (PVuV), Oryza rufipogon endornavirus (ORV), Oryza sativa endornavirus (OSV) and Vicia faba endornavirus (VFV).


   Footnotes
 
b Present address: Laboratory of Biochemistry and Genetics, NIDDK, National Institutes of Health, Bethesda, MD 20892-0830, USA.


   References
Top
Abstract
Introduction
Methods
Results and Discussion
References
 
Altschul, S. F., Madden, T. L., Schaffer, A. A., Zhang, J., Zhang, Z., Miller, W. & Lipman, D. J. (1997). Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Research 25, 3389-3402.[Abstract/Free Full Text]

Barton, G. J. (1996). Protein sequence alignment and database scanning. In Protein Structure Prediction: A Practical Approach, pp. 31-63. Edited by M. J. E. Sternberg. Oxford: IRL Press at Oxford University Press.

Barton, G. J. & Sternberg, M. J. E. (1987). A strategy for the rapid multiple alignment of protein sequences – confidence levels from tertiary structure comparisons. Journal of Molecular Biology 198, 327-337.[Medline]

Bork, P. & Gibson, T. J. (1996). Applying motif and profile searches. Methods in Enzymology 266, 162-183.[Medline]

Brown, G. G. & Finnegan, P. M. (1989). RNA plasmids. International Review of Cytology 117, 1-56.[Medline]

Dayhoff, M. O., Barker, W. C. & Hunt, L. T. (1978). A model of evolutionary change in proteins. In Atlas of Protein Sequence and Structure, vol. 5, suppl. 3. Edited by M. O. Dayhoff. Washington DC: National Biomedical Research Foundation.

Dayhoff, M. O., Barker, W. C. & Hunt, L. T. (1983). Establishing homologies in protein sequences. Methods in Enzymology 91, 524-545.[Medline]

Devereux, J., Haeberli, P. & Smithies, O. (1984). A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Research 12, 387-396.[Abstract]

Duc, G., Scalla, R. & Lefebvre, A. (1984). New developments in cytoplasmic male sterility in Vicia faba. In Vicia faba: Agronomy, Physiology and Breeding, pp. 254-260. Edited by P. D. Hebblethwaite, T. C. K. Dawkins, M. C. Heath & G. Lockwood. The Hague: Martinus Nijhoff/Dr W. Junk Publishers.

Fairbanks, D. J., Smith, S. E. & Brown, J. K. (1988). Inheritance of large mitochondrial RNAs in alfalfa. Theoretical and Applied Genetics 76, 619-622.

French, R. & Ahlquist, P. (1988). Characterisation and engineering of sequences controlling in vivo synthesis of brome mosaic virus subgenomic RNA. Journal of Virology 62, 2411-2420.[Medline]

Fukuhara, T., Moriyama, H., Pak, J. K., Hyakutake, T. & Nitta, T. (1993). Enigmatic double-stranded RNA in Japonica rice. Plant Molecular Biology 21, 1121-1130.[Medline]

Fukuhara, T., Moriyama, H. & Nitta, T. (1995). The unusual structure of a novel RNA replicon in rice. Journal of Biological Chemistry 270, 18147-18149.[Abstract/Free Full Text]

Gabriel, C. J., Walsh, R. & Nolt, B. L. (1987). Evidence for a latent virus-like agent in cassava. Phytopathology 77, 92-95.

Ghabrial, S. A., Bozarth, R. F., Buck, K. W., Yamashita, S., Martelli, G. P. & Milne, R. G. (1995). Family Partitiviridae. In Virus Taxonomy: Classification and Nomenclature of Viruses, pp. 253-260. Edited by F. A. Murphy, C. M. Fauquet, D. H. L. Bishop, S. A. Ghabrial, A. W. Jarvis, G. P. Martelli, M. A. Mayo & M. D. Summers. Wien: Springer-Verlag.

Goldbach, R. & de Haan, P. (1994). RNA viral supergroups and the evolution of RNA viruses. In The Evolutionary Biology of Viruses, pp. 105-119. Edited by S. Morse. New York: Raven Press.

Henikoff, S. & Henikof, J. G. (1992). Amino acid substitution matrices from protein blocks. Proceedings of the National Academy of Sciences, USA 89, 10915-10919.[Abstract]

Hillman, B. I., Fulbright, D. W., Nuss, D. L. & van Alfen, N. K. (1995). Family: Hypoviridae. In Virus Taxonomy: Classification and Nomenclature of Viruses, pp. 261-264. Edited by F. A. Murphy, C. M. Fauquet, D. H. L. Bishop, S. A. Ghabrial, A. W. Jarvis, G. P. Martelli, M. A. Mayo & M. D. Summers. Wien: Springer-Verlag.

Koonin, E. V. & Dolja, V. V. (1993). Evolution and taxonomy of positive-strand RNA viruses: implications of comparative analysis of amino acid sequences. Critical Reviews in Biochemistry and Molecular Biology 28, 375-430.[Abstract]

Lefebvre, A., Scalla, R. & Pfeiffer, P. (1990). The double-stranded RNA associated with the ‘447’ cytoplasmic male sterility in Vicia faba is packaged together with its replicase in cytoplasmic membranous vesicles. Plant Molecular Biology 14, 477-490.[Medline]

Mayo, M. A., Berns, K. I., Fritsch, C., Kaper, J. M., Jackson, A. O., Leibowitz, M. J. & Taylor, J. M. (1995). Subviral agents: satellites. In Virus Taxonomy: Classification and Nomenclature of Viruses, pp. 487-492. Edited by F. A. Murphy, C. M. Fauquet, D. H. L. Bishop, S. A. Ghabrial, A. W. Jarvis, G. P. Martelli, M. A. Mayo & M. D. Summers. Wien: Springer-Verlag.

Moriyama, H., Nitta, T. & Fukuhara, T. (1995). Double-stranded RNA in rice: a novel RNA replicon in plants. Molecular and General Genetics 248, 364-369.[Medline]

Moriyama, H., Kanaya, K., Wang, J. Z., Nitta, T. & Fukuhara, T. (1996). Stringently and developmentally regulated levels of a cytoplasmic double-stranded RNA and its high-efficiency transmission via egg and pollen in rice. Plant Molecular Biology 31, 713-719.[Medline]

Moriyama, H., Horiuchi, H., Koga, R. & Fukuhara, T. (1999). Molecular characterization of two endogenous double-stranded RNAs in rice and their inheritance by interspecific hybrids. Journal of Biological Chemistry 274, 6882-6888.[Abstract/Free Full Text]

Murant, A. F., Robinson, D. J. & Gibbs, M. J. (1995). Genus: Umbravirus. In Virus Taxonomy: Classification and Nomenclature of Viruses, pp. 388-391. Edited by F. A. Murphy, C. M. Fauquet, D. H. L. Bishop, S. A. Ghabrial, A. W. Jarvis, G. P. Martelli, M. A. Mayo & M. D. Summers. Wien: Springer-Verlag.

Nassuth, A. & Bol, J. F. (1983). Altered balance of the synthesis of plus- and minus-strand RNAs induced by RNAs 1 and 2 of alfalfa mosaic virus in the absence of RNA3. Virology 124, 75-85.

Pfeiffer, P. (1998). Nucleotide sequence, genetic organization and expression strategy of the double-stranded RNA associated with the ‘447’ cytoplasmic male sterility trait in Vicia faba. Journal of General Virology 79, 2349-2358.[Abstract]

Pfeiffer, P., Jung, J.-L., Heitzler, J. & Keith, G. (1993). Unusual structure of the double-stranded RNA associated with the ‘447’ cytoplasmic male sterility in Vicia faba. Journal of General Virology 74, 1167-1173.[Abstract]

Shapira, R., Choi, G. H. & Nuss, D. L. (1991). Virus-like genetic organization and expression strategy for a double-stranded RNA genetic element associated with biological control of chestnut blight. EMBO Journal 10, 731-739.[Abstract]

Strimmer, K. & von Haeseler, A. (1996). Quartet puzzling – a quartet maximum-likelihood method for reconstructing tree topologies. Molecular Biology and Evolution 13, 964-969.[Free Full Text]

Thompson, J. D., Higgins, D. G. & Gibson, T. J. (1994). CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position specific gap penalties and weight matrix choice. Nucleic Acids Research 22, 4673-4680.[Abstract]

Turpen, T., Grill, L. K. & Garger, S. J. (1988). On the mechanism of cytoplasmic male sterility in the 447 line of Vicia faba. Plant Molecular Biology 10, 489-497.

Valverde, R. A. & Fontenot, J. F. (1990). Variation in double-stranded ribonucleic acid among pepper cultivars. Journal of the American Society for Horticultural Science 116, 903-905.

Valverde, R. A., Nameth, S., Abdallha, O., Al-Musa, O., Desjardins, P. & Dodds, J. A. (1990). Indigenous double-stranded RNA from pepper (Capsicum annum). Plant Science 67, 195-201.

van der Kuyl, A. C., Neelemen, L. & Bol, J. F. (1991). Role of alfalfa mosaic virus coat protein in regulation of the balance between viral plus and minus strand RNA synthesis. Virology 185, 496-499.[Medline]

Wakarchuk, D. A. & Hamilton, R. I. (1985). Cellular double-stranded RNA in Phaseolus vulgaris. Plant Molecular Biology 5, 55-63.

Wakarchuk, D. A. & Hamilton, R. I. (1990). Partial nucleotide sequence from enigmatic dsRNAs in Phaseolus vulgaris. Plant Molecular Biology 14, 637-639.[Medline]

Zabalgogeazcoa, I. A. & Gildow, F. E. (1992). Double-stranded ribonucleic acid in ‘Barsoy’ barley. Plant Science 83, 187-194.

Zanotto, P. M. de A., Gibbs, M. J., Gould, E. A. & Holmes, E. C. (1996). A re-evaluation of the higher taxonomy of viruses based on RNA polymerases. Journal of Virology 70, 6083-6096.[Abstract]

Received 7 June 1999; accepted 10 September 1999.