Jeffrey W. Davies

The nucleotide sequence of maize streak virus DNA.

The nucleotide sequence of the DNA of maize streak virus (MSV) has been determined. The data were accommodated into one DNA circle of 2687 nucleotides, in contrast to previously characterised geminiviruses which have been shown to possess two circles of DNA. Comparison of the nucleotide sequences of the DNA of MSV with those of cassava latent virus (CLV) and tomato golden mosaic virus (TGMV) showed no detectable homology. Analysis of open reading frames revealed seven potential coding regions for proteins of mol. wt. greater than or equal to 10 000, three in the viral (+) sense and four in the complementary (‐) sense. The position of likely transcription signals on the MSV DNA sequence would suggest a bidirectional strategy of transcription as proposed for CLV and TGMV. Nine inverted repeat sequences which have a potential of forming hairpin structures of delta G greater than or equal to ‐14 kcal/mol have been detected. Three of these hairpin structures are in non‐coding regions and could be involved in the regulation of transcription and/or replication.

Specificity of Agrobacterium-mediated delivery of maize streak virus DNA to members of the Gramineae

Parameters affecting the efficiency of agroinfection of maize streak virus (MSV) in maize have been determined. Monomeric units, cloned at a number of sites in the MSV genome were not infectious but multimeric units containing partial duplications were equally as infectious as complete tandem dimeric clones. Inoculation of tandem dimeric units conjugated into different strains of Agrobacterium showed that both A. tumefaciens and A. rhizogenes were able to transfer DNA to maize and this ability was Ti (or Ri) plasmid-specific. Nopaline strains of A. tumefaciens and both agropine and mannopine A. rhizogenes strains efficiently transferred MSV DNA to maize. A number of strains were capable of MSV DNA transfer to other members of the Gramineae, providing information which may be essential for Agrobacterium-mediated transformation of monocotyledonous plants.

Geminivirus genes and vectors

The geminiviruses are very small plant viruses with circular single-stranded DNA genomes. Recent advances have identified genes involved in replication, spread of virus or DNA in the plant, and insect transmission. Gene replacement experiments suggest that useful plant gene expression vectors can be constructed from these viruses.

Aphid transmission and a polypeptide are specified by a defined region of the cauliflower mosaic virus genome

Infection of young turnip leaves with an aphid-transmissible isolate, Cabb B-JI, of cauliflower mosaic virus (CaMV) causes synthesis of an Mr 18 000 polypeptide (p18) which co-purifies with virus inclusion bodies. This polypeptide is not detectable in leaves infected with either of two aphid non-transmissible isolates. Campbell and CM4-184. Construction in vitro, of hybrid genomes between Cabb B-JI and Campbell isolates demonstrates that aphid transmissibility and presence of p18 is dependent on the small genome fragment from the BstEII site to the XhoI site. A deletion made in this fragment within open reading frame (ORF) II causes loss of aphid transmissibility and also terminates production of p18. We conclude that aphid transmissibility and the presence of p18 are related to the expression of ORF II of the CaMV genome.

Rice tungro bacilliform virus DNA independently infects rice after Agrobacterium-mediated transfer

In nature, rice tungro disease is caused by an RNA and a DNA virus complex, but we have obtained an independently infectious clone of rice tungro bacilliform virus (RTBV) DNA. Infectivity could be demonstrated only when a more than unit-length copy was cloned in the Agrobacterium binary vector Bin 19 and agroinoculated into rice plants. Rice plants thus agroinfected with cloned RTBV DNA showed typical symptoms of tungro disease, presence of viral DNA and bacilliform particles, and could be used as a source of virus to infect healthy plants by the green leafhopper (Nephotettix virescens). The importance of this infectious clone in understanding the molecular biology of RTBV and the rice tungro disease is discussed.

Approaches to nonconventional control of plant virus diseases

Abstract Plant viruses cause significant losses to crops worldwide and the three basic approaches to controlling them have not been overly successful. The new concept of nonconventional resistance, which involves transforming plants with nucleic acid sequences that interfere with the viral infection cycle, is a promising new approach. The current nonconventional strategies, those of coat protein‐mediated protection, antisense nucleic acids, satellite sequences, defective interfering molecules, and nonstructural genes, are reviewed with their advantages and disadvantages being discussed. The question of what exactly is nonconventional resistance is raised and suggestions are made for defining resistance. A range of strategies that are likely to be developed in the future are outlined together with some guiding principles for selection and deployment of these forms of resistance.

The nucleotide sequence of a geminivirus from Digitaria sanguinalis

The encapsidated single-stranded circular DNA of a geminivirus isolated from Digitaria sanguinalis has been sequenced. The data obtained are consistent with there being one DNA circle of 2701 nucleotides. Comparison of the nucleotide sequence with those of maize streak virus (MSV) and wheat dwarf virus showed 64 and 47% DNA homology, respectively. The sequence has four potential coding regions for proteins of greater than 10 kDa, two in the viral (+) sense and two in the complementary (-) sense. Each of these potential coding regions has a highly homologous counterpart among the seven open reading frames previously described for MSV. Virion DNA contained, in addition to the circular single-stranded DNA, a population of small DNA molecules similar to those associated with MSV particles. A comparison with MSV DNA of the region complementary to these small DNA molecules revealed conserved sequences, which may have a role in defining the limits of these primer-like molecules.

A putative primer for second-strand DNA synthesis of maize streak virus is virion-associated.

We have isolated, from maize streak virus (MSV) preparations, a population of ‘nested’ DNA molecules. These molecules have ribonucleotides covalently linked to the DNA species’ discrete 5′ deoxyribonucleotide terminus. The major species has a DNA sequence of 80 nucleotides which is complementary to a region 5′ of two hairpin structures on the MSV genome, almost exclusively in an intergenic region. These molecules have been used to prime the synthesis in vitro of a complementary strand to virion DNA, initiating this reaction at one site on the genome.

Location and sequence of a region of cauliflower mosaic virus gene 2 responsible for aphid transmissibility

Abstract The nucleotide sequence of the aphid nontransmissible Campbell isolate of cauliflower mosaic virus has been determined between the BstEll and Xho I sites, the region known to be associated with aphid transmission. Comparison of the DNA sequence with that of the aphid transmissible isolates revealed no gross structural alterations in any of the open reading frames (ORFs). Recombinant DNA hybrid constructs and in vitro alteration of ORF VII were used to verify that ORF VII was not involved in the regulation of the downstream ORF II. Instead, a modification within a small region of ORF II which alters only one amino acid of the potential protein product relative to the aphid-transmissible isolate was found to be responsible for the lack of aphid transmission in the Campbell isolate.

Agrobacterium-mediated infectivity of cloned digitaria streak virus DNA

A monomeric clone of double-stranded DNA synthesized in vitro DNA of the geminivirus Digitaria streak (DSV) was subcloned as a tandem dimeric unit into a binary vector of Agrobacterium tumefaciens, creating a plasmid pDS2. Inoculation of digitaria sanguinalis with A. tumefaciens carrying pDS2 resulted in viral infection. The symptoms, virus particles, and DNA forms obtained were indistinguishable from those of a natural DSV infection of D. sanguinalis. Inoculations have also induced infections in Zea mays and Avena sativa. The sequence of the Agrobacterium-mediated infectious clone of DSV has been determined.