Evolutionary relationship of alfalfa mosaic virus with cucumber mosaic virus and brome mosaic virus

The amino acid sequences of the non-structural protein (molecular weight 35,000; 3a protein) from three plant viruses — cucumber mosaic, brome mosaic and alfalfa mosaic have been systematically compared using the partial genomic sequences for these three viruses already available. The 3a protein of cucumber mosaic virus has an amino acid sequence homology of 33.7% with the corresponding protein of brome mosaic virus. A similar protein from alfalfa mosaic virus has a homology of 18.2% and 14.2% with the protein from brome mosaic virus and cucumber mosaic virus, respectively. These results suggest that the three plant viruses are evolutionarily related, although, the evolutionary distance between alfalfa mosaic virus and cucumber mosaic virus or brome mosaic virus is much larger than the corresponding distance between the latter two viruses.     

Similarities in the genomic sequence and coat protein structure of plant virsuses

The genomic sequences of several RNA plant viruses including cucumber mosaic virus, brome mosaic virus, alfalfa mosaic virus and tobacco mosaic virus have become available recently. The former two viruses are icosahedral while the latter two are bullet and rod shaped, respectively in particle morphology. The non-structural 3a proteins of cucumber mosaic virus and brome mosaic virus have an amino acid sequence homology of 35% and hence are evolutionarily related. In contrast, the coat proteins exhibit little homology, although the circular dichroism spectrum of these viruses are similar. The non-coding regions of the genome also exhibit variable but extensive homology. Comparison of the brome mosaic virus and alfalfa mosaic virus sequences reveals that they are probably related although with a much larger evolutionary distance. The polypeptide folds of the coat protein of three biologically distinct isometric plant viruses, tomato Bushy stunt virus, southern bean mosaic virus and satellite tobacco necrosis virus have been shown to display a striking resemblance. All of them consist of a topologically similar 8-standard β-Barrel. The implications of these studies to the understanding of the evolution of plant viruses will be discussed.     

The complete nucleotide sequence of the potexvirus white clover mosaic virus.

The complete nucleotide sequence (5845 nucleotides) of the genomic RNA of the potexvirus white clover mosaic virus (WC1MV) has been determined from a set of overlapping cDNA clones. Forty of the most 5'-terminal nucleotides of WC1MV showed homology to the 5' sequences of other potexviruses. The genome contained five open reading frames which coded for proteins of Mr 147, 417, Mr 26,356, Mr 12,989, Mr 7,219 and Mr 20,684 (the coat protein). The Mr 147,417 protein had domains of amino acid sequence homology with putative polymerases of other RNA viruses. The Mr 26,356 and Mr 12,989 proteins had homology with proteins of the hordeivirus barley stripe mosaic virus RNA beta and the furovirus beet necrotic yellow vein virus (BNYVV) RNA-2. A portion of the Mr 26,356 protein was also conserved in the cylindrical inclusion proteins of two potyviruses. The Mr 7,219 protein had homology with the 25K putative fungal transmission factor of BNYVV RNA-3.     

Homologous sequences in non-structural proteins from cowpea mosaic virus and picornaviruses

Computer analyses have revealed sequence homology between two non-structural proteins encoded by cowpea mosaic virus (CPMV), and corresponding proteins encoded by two picornaviruses, poliovirus and foot-and-mouth disease virus. A region of 535 amino acids in the 87-K polypeptide from CPMV was found to be homologous to the RNA-dependent RNA polymerases from both picornaviruses, the best matches being found where the picornaviral proteins most resemble each other. Additionally, the 58-K polypeptide from CPMV and polypeptide P2-X from poliovirus contain a conserved region of 143 amino acids. Based on the homology observed, a genetic map of the CPMV genome has been constructed in which the 87-K polypeptide represents the core polymerase domain of the CPMV replicase. These results have implications for the evolution of RNA viruses, and mechanisms are discussed which may explain the existence of homology between picornaviruses (animal viruses with single genomic RNAs) and comoviruses (plant viruses with two genomic RNAs).     

Identification and study of tobacco mosaic virus movement function by complementation tests

The phenomenon of trans-complementation of cell-to-cell movement between plant positive-strand RNA viruses is discussed with an emphasis on tobamoviruses. Attention is focused on complementation between tobamoviruses (coding for a single movement protein, MP) and two groups of viruses that contain the triple block of MP genes and require four (potato virus X) or three (barley stripe mosaic virus) proteins for cell-to-cell movement. The highlights of complementation data obtained by different experimental approaches are given, including (i) double infections with movement-deficient (dependent) and helper viruses; (ii) infections with recombinant viral genomes bearing a heterologous MP gene; (iii) complementation of a movement-deficient virus in transgenic plants expressing the MP of a helper virus; and (iv) co-bombardment of plant tissues with the cDNAs of a movement-dependent virus genome and the MP gene of a helper virus.     

Chimeras between Oilseed rape mosaic virus and Tobacco mosaic virus highlight the relevant role of the tobamoviral RdRp as pathogenicity determinant in several hosts

Oilseed rape mosaic virus (ORMV) is a tobamovirus taxonomically distinct from the type member of the genus, Tobacco mosaic virus (TMV). Both viruses display a specific host range, although they share certain hosts, such as Arabidopsis thaliana , Nicotiana benthamiana and N. tabacum , on which they induce different symptoms. Using a gain-of-symptom approach, we generated chimeric viruses, starting from a TMV infectious clone, over which different regions of ORMV were exchanged with their corresponding regions in the TMV genome. This approach allowed the association of pathogenicity determinants to certain genes within the ORMV genome. A general trend was observed associating the viral origin of the RNA-dependent RNA-polymerase ( RdRp ) gene and the gain of symptoms. In A. thaliana and N. benthamiana , chimeric viruses were unable to reproduce the symptoms induced by the parental viruses, leading to disease states which could be described as intermediate, and variable in some cases. In contrast, a hypersensitive reaction caused by both of these viruses on N -gene-bearing tobaccos could be found in resistance reactions to all chimeric viruses, suggesting that the avirulence determinant maps similarly in both viruses. A systemic necrotic spotting typical of non- N -gene tobaccos infected with ORMV was associated with the polymerase domain of RdRp . To our knowledge, this is the first time that this controversial portion of the tobamovirus genome has been identified directly as a pathogenicity determinant. None of the reactions of the chimeric viruses could be correlated with increases or decreases in virus titres in the infections.     

Nucleotide sequence of the brome mosaic virus genome and its implications for viral replication

The nucleotide sequences of brome mosaic virus (BMV) RNAs 1 (3234 bases) and 2 (2865 bases) have been determined, completing the primary structure of the 8200 base tripartite BMV genome. cDNA clones covering 99% of BMV RNA1 and a full-length cDNA clone of BMV RNA2 were isolated in the course of this work. Extensive sequence homology and known interaction with several proteins suggest that the 3' ends of the BMV RNAs are the major regulatory regions of the genome. Smaller regions at the 5' ends of RNAs 1 and 2 show strong homology to each other and lesser homology to RNA3. These and other features of the sequences are discussed in relation to replication, regulation and evolution of the BMV genome.     

Cucumber mosaic virus genome is encapsidated in alfalfa mosaic virus coat protein expressed in transgenic tobacco plants

The expression of viral coat protein (CP) in transgenic plants has been shown to be very effective in virus plant protection. However, the introduction of CP genes into plants presents the potential risk of the encapsidation of a superinfecting viral genome in the transgenic protein, an event which could change the epidemiology of the disease. To detect the potential heterologous encapsidation of the cucumber mosaic virus (CMV) genome by alfalfa mosaic virus (AIMV) CP expressed in transgenic tobacco plants, a system of immunocapture (IC) and amplification by polymerase chain reaction (PCR) was optimized. This provided high sensitivity and reliable selection of the heterologously encapsidated CMV genome in the presence of natural CMV particles. As little as 2 pg of virus could be detected by immunocapture/polymerase chain reaction (IC/PCR) technique. Evidence for heterologous encapsidation of the CMV genome was found in 11 of the 33 transgenic plants tested two weeks after CMV inoculation. This demonstrates a significant rate of heterologous encapsidation events between two unrelated viruses in transgenic plants. Since CP is involved in the interactions of the virus particle with its vector, the release in the field of such transgenic plants could alter the transmission properties of some important viruses.     

Homology between the proteins encoded by tobacco mosaic virus and two tricornaviruses

Summary A comparison was made of the amino acid sequences of the proteins encoded by RNAs 1 and 2 of alfalfa mosaic virus (A1MV) and brome mosaic virus (BMV), and the 126K and 183K proteins encoded by tobacco mosaic virus (TMV). Three blocks of extensive homology of about 200 to 350 amino acids each were observed. Two of these blocks are located in the A1MV and BMV RNA 1 encoded proteins and the TMV encoded 126K protein; they are situated at the N-terminus and C-terminus, respectively. The third block is located in the A1MV and BMV RNA 2 encoded proteins and the C-terminal part of the TMV encoded 183K protein. These homologies are discussed with respect to the functional equivalence of these putative replicase proteins and a possible evolutionary connection between A1MV, BMV and TMV.     

Sequence homology among the coat proteins of gemini viruses

The coat proteins of the gemini viruses - African cassava latent virus, tomato golden mosaic virus and maize streak virus - are shown to have reasonable to good amino acid sequence homology. It is suggested that the maize streak virus genome is ancestral and the bipartite genomes of the other viruses evolved from it.     

Tobacco mosaic virus and the study of early events in virus infections

In order to establish infections, viruses must be delivered to the cells of potential hosts and must then engage in activities that enable their genomes to be expressed and replicated. With most viruses, the events that precede the onset of production of progeny virus particles are referred to as the early events and, in the case of positive-strand RNA viruses, they include the initial interaction with and entry of host cells and the release (uncoating) of the genome from the virus particles. Though the early events remain one of the more poorly understood areas of plant virology, the virus with which most of the relevant research has been performed is tobacco mosaic virus (TMV). In spite of this effort, there remains much uncertainty about the form or constituent of the virus that actually enters the initially invaded cell in a plant and about the mechanism(s) that trigger the subsequent uncoating (virion disassembly) reactions. A variety of approaches have been used in attempts to determine the fate of TMV particles that are involved in the establishment of an infection and these are briefly described in this review. In some recent work, it has been proposed that the uncoating process involves the bidirectional release of coat protein subunits from the viral RNA and that these activities may be mediated by cotranslational and coreplicational disassembly mechanisms.     

The complete sequence of a sugarcane mosaic virus isolate causing maize dwarf mosaic disease in China

The complete sequence of a potyvirus from maize in Zhejiang Province was determined. The RNA was 9596 nucleotides long, excluding the 3'-poly (A) tail, and there was a single long open reading frame (ORF) of 9192 nts encoding a 346.1 ku polyprotein. The polyprotein had substantial amino acid sequence homology with those encoded by the RNAs of a Chinese isolate of sorghum mosaic virus (SrMV-C) and a Bulgarian isolate of maize dwarf mosaic virus, but it was most closely related to sugarcane mosaic virus (SCMV) isolates, for which only partial sequences have been published. According to the published criteria for distinguishing potyviruses, the sequence reported here is clearly a strain of SCMV, but it also showed a surprisingly high amino acid homology with SrMV-C in the HC-Pro, P3 and Cl proteins.     

The complete sequence of a sugarcane mosaic virus isolate causing maize dwarf mosaic disease in China

The complete sequence of a potyvirus from maize in Zhejiang Province was determined. The RNA was 9596 nucleotides long, excluding the 3′-poly (A) tail, and there was a single long open reading frame (ORF) of 9192 nts encoding a 346.1 ku polyprotein. The polyprotein had substantial amino acid sequence homology with those encoded by the RNAs of a Chinese isolate of sorghum mosaic virus (SrMV-C) and a Bulgarian isolate of maize dwarf mosaic virus, but it was most closely related to sugarcane mosaic virus (SCMV) isolates, for which only partial sequences have been published. According to the published criteria for distinguishing potyviruses, the sequence reported here is clearly a strain of SCMV, but it also showed a surprisingly high amino acid homology with SrMV-C in the HC-Pro, P3 and Cl proteins.     

The sequence of carnation etched ring virus DNA: comparison with cauliflower mosaic virus and retroviruses

Carnation etched ring virus (CERV) DNA comprises 7932 bp. CERV primer binding sites and overall genome organization are similar to those of the related cauliflower mosaic virus (CaMV). The six open reading frames of CERV showed amino acid homology (50-80%) with CaMV ORFs I-VI; no homologues of CaMV ORFs VII or VIII were found. CERV ORFs 1-5 interface each other with the sequence ATGA. The comparison of CERV ORF5 with CaMV ORFV highlighted regions which show homologies to retrovirus gag/pol protease, RNase H and DNA polymerase domains; the possibility that the DNA polymerase domain comprises two subdomains, operating off different templates, is discussed. Both CERV and CaMV ORFs I have sequence homology to tobacco mosaic virus P30 and plastocyanin.     

Sindbis virus proteins nsP1 and nsP2 contain homology to nonstructural proteins from several RNA plant viruses.

Although the genetic organization of tobacco mosaic virus (TMV) differs considerably from that of the tripartite viruses (alfalfa mosaic virus [AlMV] and brome mosaic virus [BMV]), all of these RNA plant viruses share three domains of homology among their nonstructural proteins. One such domain, common to the AlMV and BMV 2a proteins and the readthrough portion of TMV p183, is also homologous to the readthrough protein nsP4 of Sindbis virus (Haseloff et al., Proc. Natl. Acad. Sci. U.S.A. 81:4358-4362, 1984). Two more domains are conserved among the AlMV and BMV 1a proteins and TMV p126. We show here that these domains have homology with portions of the Sindbis proteins nsP1 and nsP2, respectively. These results strengthen the view that the four viruses share mechanistic similarities in their replication strategies and may be evolutionarily related. These results also suggest that either the AlMV 1a, BMV 1a, and TMV p126 proteins are multifunctional or Sindbis proteins nsP1 and nsP2 function together as subunits in a single complex.     

Elucidation of the genome organization of tobacco mosaic virus

Proteins unique to tobacco mosaic virus (TMV)-infected plants were detected in the 1970s by electrophoretic analyses of extracts of virus-infected tissues, comparing their proteins to those generated in extracts of uninfected tissues. The genome organization of TMV was deduced principally from studies involving in vitro translation of proteins from the genomic and subgenomic messenger RNAs. The ultimate analysis of the TMV genome came in 1982 when P. Goelet and colleagues sequenced the entire genome. Studies leading to the elucidation of the TMV genome organization are described below.     

Comparison of barley stripe mosaic virus strains

BSMV (barley stripe mosaic virus) particles were obtained in a pure state from infected host plant tissues of Hordeum vulgare. The three genomic parities (alpha, beta and gamma) were amplified by PCR using specific primers for each particle; each was cloned. Partial sequence of the alpha, beta and gamma segments was determined for the Egyptian isolate of barley stripe mosaic virus (BSMV AE1). Alignment of nucleotide sequences with that of other known strains of the virus, BSMV type strains (CV17, ND18 and China), and the generation of phylogenetic trees was performed. A low level of homology was detected comparing 467 bp of the a and 643 bp of the segments to that of the other strains, and thus BSMV alpha and beta segments were in separate clusters. However, 1154 bp of the gamma segments of BSMV AE1 showed a high level of homology especially to strain BSMV ND18, as they both formed a distinct cluster. Northern blotting of pure BSMV AE1 virus and H. vulgare-infected tissue were compared using an alpha ND18 specific probe. Western blotting using antibodies specific for the coat protein (CP) and the triple gene block 1 (TGB1) protein, which are both encoded by the beta ND18 segment, still indicated a high level of similarity between proteins produced by BSMV ND18 and AE1. We suggest that the BSMV AE1 isolate is a distinct strain of BSMV which reflects the genetic evolutionary divergence among BSMV strains and members of the Hordeivirus group.     

Characterization of cymbidium mosaic virus coat protein gene and its expression in transgenic tobacco plants

Cymbidium mosaic virus (CyMV) is the most prevalent virus infecting orchids. Here, we report the isolation of partial cDNA clones encoding the genomic RNA of CyMV. Like most of the polyadenylated monopartite positive-strand RNA viruses, the open reading frame (ORF) coding for the viral coat protein (CP) is located at the 3 end. The ORF predicts a polypeptide chain of 220 amino acids with a molecular weight of 23 600. Sequence comparison of this ORF to the CP sequences of potato virus X(PVX) and white clover mosaic virus (WCIMV) revealed a strong amino acid homology in the mid-portion of the CP, but the overall homology was low. The CyMV CP gene was placed downstream of a cauliflower mosaic virus 35S promoter and the chimaeric gene was transferred into Nicotiana benthamiana. Transgenic plants expressing the CyMV CP were protected against CyMV infection.