Biomedical subjects

M A Rezaian

Publications and source records attributed to M A Rezaian.

At least 19 recordsLinked to original sources

PubMed1998

Efficient cloning of cDNA from grapevine leafroll-associated virus 4 and demonstration of probe specificity by the viral antibody.

Using a random-PCR method, a cDNA clone (LR4) was constructed from the replicative form dsRNA of grapevine leafroll-associated virus 4 (GLRaV-4). Northern blot analysis showed hybridization of LR4 to dsRNA in an extract of a Thompson Seedless grapevine clone from which GLRaV-4 was isolated originally by Hu et al. (1990). The cDNA clone was sequenced and shown to be specific to GLRaV-4 by reverse-transcription-PCR using GLRaV-4 particles enriched by the virus antibody coupled to magnetic beads. Reverse-transcription-PCR was used successfully to screen different varieties of grapevines for the virus. Western blot analysis of GLRaV-4 extracts from different varieties of infected grapevines revealed two distinct species of capsid protein with estimated Mr of either 35500 or 38000 depending on the variety used. Both proteins reacted with polyclonal as well as monoclonal antibodies.

Antibodies, Viral↗

PubMed1997

A novel subviral agent associated with a geminivirus: the first report of a DNA satellite.

Numerous plant RNA viruses have associated with them satellite (sat) RNAs that have little or no nucleotide sequence similarity to either the viral or host genomes but are completely dependent on the helper virus for replication. We report here on the discovery of a 682-nt circular DNA satellite associated with tomato leaf curl geminivirus (TLCV) infection in northern Australia. This is the first demonstration that satellite molecules are not limited to RNA viral systems. The DNA satellite (TLCV sat-DNA) is strictly dependent for replication on the helper virus replication-associated protein and is encapsidated by TLCV coat protein. It has no significant open reading frames, and it shows no significant sequence similarity to the 2766-nt helper-virus genome except for two short motifs present in separate putative stem-loop structures: TAATATTAC, which is universally conserved in all geminiviruses, and AATCGGTGTC, which is identical to a putative replication-associated protein binding motif in TLCV. Replication of TLCV sat-DNA is also supported by other taxonomically distinct geminiviruses, including tomato yellow leaf curl virus, African cassava mosaic virus, and beet curly top virus. Therefore, this unique DNA satellite does not appear to strictly conform with the requirements that dictate the specificity of interaction of geminiviral replication-associated proteins with their cognate origins as predicted by the current model of geminivirus replication.

Base Sequence↗

PubMed1996

Plant virus DNA replication processes in Agrobacterium: insight into the origins of geminiviruses?

Agrobacterium tumefaciens, a bacterial plant pathogen, when transformed with plasmid constructs containing greater than unit length DNA of tomato leaf curl geminivirus accumulates viral replicative form DNAs indistinguishable from those produced in infected plants. The accumulation of the viral DNA species depends on the presence of two origins of replication in the DNA constructs and is drastically reduced by introducing mutations into the viral replication-associated protein (Rep or C1) ORF, indicating that an active viral replication process is occurring in the bacterial cell. The accumulation of these viral DNA species is not affected by mutations or deletions in the other viral open reading frames. The observation that geminivirus DNA replication functions are supported by the bacterial cellular machinery provides evidence for the theory that these circular single-stranded DNA viruses have evolved from prokaryotic episomal replicons.

Biological Evolution↗

PubMed1995

The termini of a new citrus viroid contain duplications of the central conserved regions from two viroid groups.

A citrus viroid associated with dwarfing, CVdIIIA, has been sequenced and its 294 nucleotide residues can be arranged to form the typical rod-like secondary structure of other viroids with 71% of nucleotides base-paired. CVdIIIA has greatest sequence similarity with apple scar skin viroid (ASSVd; 69%) and has the central sequence which is conserved in the ASSVd group. CVdIIIA is the smallest member of the ASSVd group but contains the terminal conserved region shared by all viroids over 300 nucleotides. The two ends of CVdIIIA are highly unusual in that each end appears to be derived from the conserved central core region of a different viroid group.

Base Sequence↗

PubMed1994

ORF C4 of tomato leaf curl geminivirus is a determinant of symptom severity.

A tomato leaf curl virus (TLCV) mutant has been constructed in vitro that contains T-to-C replacements at nucleotides 2457 and 2463 within the C4 open reading frame (ORF). The mutations destroy the two possible initiator AUG codons for the C4 ORF without disrupting the coding capacity of the C1 ORF which entirely overlaps the C4 ORF. Agroinoculation of the C4 mutant TLCV into three alternative experimental hosts for the virus (Datura stramonium, Lycopersicon esculentum, and Nicotiana tabacum) gives rise to infections which show dramatically reduced symptoms when compared to a wild-type infection, while retaining wild-type levels of all viral DNA species. In most cases the mutations were stably inherited by progeny virus. However, a single tomato plant inoculated with the mutant developed phenotypically wild-type symptoms and was subsequently shown to contain progeny virus in which the mutation at position 2457 had reverted to wild-type sequence, indicating that this AUG may be the site of initiation of translation of the C4 product in the wild-type virus. The results suggest that the C4 ORF encodes a polypeptide which is not required by TLCV to replicate or to spread through the host plant, but is involved in symptom development.

Base Sequence↗

PubMed1993

Mapping of the polycistronic RNAs of tomato leaf curl geminivirus.

Four major transcripts from infected tomato were identified and mapped onto the monopartite genome of the geminivirus tomato leaf curl virus (TLCV). The C1, C2, and C3 ORFs are spanned by one transcript and a second internal RNA covered C2 and C3. Both these RNAs have their counterparts in the DNA A components of the bipartite subgroup of geminiviruses. The 5' ends of the virion-sense RNAs map either side of the first in-frame AUG of the V1 ORF. The 3' ends of the virion-sense RNAs are coterminal and overlap with the 3' ends of the complementary-sense RNAs. All of the RNAs have transcription regulatory sequences close to their mapped termini and the presence of overlapping transcripts suggests that temporal regulation of their synthesis may occur. The translation of these polycistronic RNAs is discussed in the light of the RNA mapping data.

Amino Acid Sequence↗

PubMed1993

Mutagenesis of the virion-sense open reading frames of tomato leaf curl geminivirus.

A series of frame shift, deletion, and inversion mutants in the virion-sense open reading frames (ORFs) of the monopartite geminivirus tomato leaf curl virus have been constructed and their ability to replicate, produce single-stranded DNA, spread, and cause symptoms in tomato plants has been investigated. Disruptions in the V1 ORF lead to symptomless, systemic infections with a reduced titer of all viral DNA forms while interruptions in the V2 (coat protein) ORF disrupted spread of the virus. Mutagenesis of the virion-sense ORFs did not affect the replication of viral double-stranded DNA, although both V1 and V2 products appear to play a role in the accumulation of viral single-stranded DNA.

Blotting, Southern↗

PubMed1993

Analysis of sequence variation in grapevine yellow speckle viroid 1 reveals two distinct alternative structures for the pathogenic domain.

The nucleotide sequences of 24 full-length cDNA clones prepared from a field isolate of grapevine yellow speckle viroid 1 (GYSV 1) have been determined and compared with that of the previously published GYSV 1 sequence. A large number of sequence variations were observed, the majority of which occurred in the pathogenicity domain (P domain) of the viroid. The GYSV 1 variants could be divided into two types each containing a distinct secondary structure within the P domain. Monomeric exact-length RNA transcripts produced from individual clones belonging to both types were infectious in viroid-free grapevines and produced homogenous populations of progeny viroid in vivo.

Base Sequence↗

PubMed1993

Nucleotide sequence and genome organization of tomato leaf curl geminivirus.

The genome of tomato leaf curl virus (TLCV) from Australia was cloned and its complete nucleotide sequence determined. It is a single circular ssDNA of 2766 nucleotides containing the consensus nonanucleotide sequence present in all geminiviruses. It has six open reading frames with an organization resembling that of certain other dicotyledonous plant-infecting monopartite geminiviruses, i.e. tomato yellow leaf curl and beet curly top viruses. The regulatory sequences present indicate a bidirectional mode of transcription. A dimeric TLCV DNA clone was constructed in a binary vector and used to agroinoculate three different host species. Typical virus infections were produced, confirming that the single DNA component is sufficient for infectivity.

Amino Acid Sequence↗

PubMed1992

In vitro synthesis of an infectious viroid: analysis of the infectivity of monomeric linear CEV.

Infectious monomers of citrus exocortis viroid (CEV) were synthesized in vitro precisely to predetermined sequences in microgram quantities without resorting to cloning procedures. Amplification of CEV double-stranded cDNAs fused with a T7 RNA polymerase promoter was followed by transcription of the DNA resulting in the production of an infectious linear CEV monomer. This is the first demonstration of an infectious unit length viroid synthesized in vitro. Transcripts containing 3'-OH terminal groups were infectious, demonstrating that a 2',3'-cyclic phosphate terminus is not a prerequisite for viroid infectivity as previously suggested. Conversion of the 5'-triphosphate terminus to either 5'-monophosphate or 5'-OH had little effect on infectivity. The linear RNA could be circularized using T4 RNA ligase to produce an authentic CEV molecule. This procedure, which results in the production of biologically active RNA, would allow routine application of oligonucleotide-directed mutagenesis to the study of viroids and other circular RNAs. It would also enable the in vitro synthesis and mutagenesis of infectious viral RNAs containing a 5'-G residue.

Base Sequence↗

PubMed1992

Common identity of grapevine viroids from USA and Australia revealed by PCR analysis.

Pairs of viroid-specific oligonucleotide primers were selected and used in separate reverse transcription reactions coupled with the polymerase chain reaction to obtain DNA products of predetermined sizes characteristic of each viroid. The reaction conditions allowed efficient incorporation of small amounts of 32P-dATP which enabled rapid detection of the products in polyacrylamide gels. Using this method as well as probe hybridization, the presence of grapevine yellow speckle viroids 1 and 2 (previously known as GV1B) in grapevine samples from California was demonstrated, and it was established that the Australian grapevine viroid occurs in California. These comparisons provide the basis for uniform nomenclature of grapevine viroids found in different geographical regions.

Australia↗

PubMed1990

Australian grapevine viroid--evidence for extensive recombination between viroids.

Australian grapevine viroid (AGV, 369 residues) is a novel viroid with less than 50% sequence similarity with any known viroid. Nevertheless its entire sequence can be divided into regions, each with a high sequence similarity with segments from one of citrus exocortis, potato spindle tuber, apple scar skin, and grapevine yellow speckle viroids. AGV contains the entire central conserved region of the apple scar skin viroid group and is proposed as a member of this group. AGV appears to have originated from extensive RNA recombination involving other viroids. The vegetatively propagated grapevines which have been exposed to multiple viroid infections during their long history of cultivation may have allowed such recombination.

Base Sequence↗

PubMed1989

Grapevine viroid 1B, a new member of the apple scar skin viroid group contains the left terminal region of tomato planta macho viroid.

GV1B is one of five viroids that have recently been purified from grapevines. GV1B has now been sequenced and its 363 nucleotide residues can potentially form the typical rod-like structure of viroids with 67% of nucleotides base-paired. GV1B has highest sequence similarity with grapevine yellow speckle viroid (GYSV; 73%) and has a central sequence which is conserved in GYSV and apple scar skin viroid (ASSV) which have been reported to constitute the ASSV group. Therefore, we have placed GV1B into the ASSV group. GV1B contains a direct repeat sequence at the terminal portions of its T1 and T2 regions. GV1B also contains a sequence of 69 nucleotides in the terminal portion of its T1 region which is almost identical to the corresponding region in tomato planta macho viroid (TPMV). This provides further evidence of the importance of RNA recombination in viroid evolution.

Base Sequence↗

PubMed1989

Two related viroids cause grapevine yellow speckle disease independently.

We have confirmed that two closely related circular RNA molecules previously named grapevine yellow speckel viroid (GYSV) and grapevine viroid 1B (GV1B) are indeed viroids. Electron microscopy after spreading under non-denaturing conditions revealed that GYSV has a rod-like structure typical of viroids. Purified GYSV and GV1B replicated independently in inoculated grapevine seedlings and some of the infected plants developed yellow speckle symptoms indicating that both viroids can cause grapevine yellow speckle disease. Plus-sense RNA transcripts derived from a dimeric GYSV cDNA clone induced yellow speckle symptoms in a grapevine seedling confirming the role of GYSV in the yellow speckle disease. Two oligonucleotide probes were synthesized for the detection of the two related viroids. The probes which could detect each viroid individually were used to assess correlations between the occurrence of these viroids and the incidence of the disease.

Base Sequence↗

PubMed1989

A scheme for viroid classification.

A scheme for viroid classification is proposed based on the nature of the strictly conserved core sequence present in the central portion of the secondary structure of viroids. In this scheme, all of the known viroids can be classified as potato spindle tuber-type viroids consisting of two viroid groups and avocado sunblotch-type viroids containing one viroid group.

Base Sequence↗

PubMed1988

Grapevine yellow speckle viroid: structural features of a new viroid group.

A single stranded circular RNA was isolated from grapevines infected with yellow speckle disease. The RNA which we have called grapevine yellow speckle viroid (GYSV), contains 367 nucleotide residues and has the potential to form the rod-like secondary structure characteristic of viroids. GYSV has 37% sequence homology with the recently described apple scar skin viroid (ASSV; 330 residues) and has some sequence homology with the viroids in the potato spindle tuber viroid (PSTV) group. The sequence of GYSV has characteristics which fit the structural domains described for the PSTV group. However, GYSV lacks the PSTV central conserved sequence. Instead, there is a conserved sequence in the central region of GYSV and ASSV which has the potential to form a stem loop configuration and a stable palindromic structure as does the central conserved region of the PSTV group. These structural features suggest there is a different central conserved region for GYSV and ASSV. The results support the viroid nature of GYSV and its inclusion into a separate viroid group which we suggest should be represented by ASSV.

Base Sequence↗

PubMed1988

Isolation of three viroids and a circular RNA from grapevines.

Analysis of nucleic acids from grapevine tissues by two-dimensional gel electrophoresis demonstrated the presence of two bands of circular RNA. The smaller RNA contained about 300 nucleotide residues and was identified as hop stunt viroid by nucleotide sequencing. The larger RNA band was a mixture of species and contained similar amounts of two components, referred to as RNA 1a and RNA 1b, and in addition a trace amount of citrus exocortis viroid (CEV) which became detectable only after inoculation of the mixture to tomato. The identity of CEV was determined by probe hybridization and nucleotide sequencing. Both RNAs 1a and 1b are distinct from CEV and have estimated sizes larger than those of CEV and other viroids reported so far. RNA 1a preparations were infectious in cucumber and in tomato and the recovered viroid had unique properties. We have provisionally named this viroid Australian grapevine viroid. Evidence for the autonomous replication of RNA 1b was not obtained.

Base Sequence↗

PubMed1987

Nucleic acid extraction and virus detection in grapevine.

Three extraction media for the isolation of nucleic acids from grapevines, a tissue high in polyphenols and other materials that interfere with nucleic acid extraction, were compared. When phenol was present in the initial extraction media only a small yield of soluble RNA and no high molecular weight rRNA was obtained. In the absence of phenol in conventional salt and detergent-based extraction media, rRNAs were extracted, but a major proportion of the RNAs were broken down. Using Na-perchlorate, a chaotropic salt, in a rapid procedure, it was possible to extract both high and low molecular weight RNA efficiently. This procedure enabled the detection of viral RNA, which could not be detected following phenol extraction, at the picogram levels, by dot-blot hybridization.

Nucleic Acid Hybridization↗