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At least 19 recordsLinked to original sources
PubMed1995

Molecular analysis of the essential and nonessential genetic elements in the genome of peanut chlorotic streak caulimovirus.

The DNA genome of caulimoviruses contains a set of essential genes: I (movement gene), IV (major capsid protein gene), V (reverse transcriptase gene), and VI (gene coding for a post-transcriptional activator of the expression of other virus genes). In peanut chlorotic streak caulimovirus (PCISV), three ORFs, A, B, and C, are located between genes I and IV. They are dissimilar to other caulimovirus ORFs. ORF VII of PCISV is a homolog of ORF VII of soybean chlorotic mottle caulimovirus (SoCMV), but is not similar to the nonconserved ORF VII in other caulimoviruses. The sequence complementary to a portion of tRNA(Met), thought to be essential for the priming of minus-strand DNA synthesis in caulimoviruses, is located within the coding sequence of ORF A. To explore the functional significance of ORFs VII, A, B, and C, various mutations were engineered into an infectious DNA clone of PCISV. ORFs VII and B are shown to be dispensable, while ORFs A and C are essential. ORF C is a possible functional equivalent of gene III in other caulimoviruses. Sequences within ORF A that are required for efficient priming of minus-strand synthesis are likely to extend beyond the 12-bp tRNA-binding site. Complete deletion of ORF VII was correlated with severe symptoms, notably with the necrosis of apical meristems. Significance of these observations for the understanding of replication and pathogenesis of plant pararetroviruses and for the improvement of caulimovirus-based expression vectors is discussed.

Arachis↗
PubMed2026

The complete genomic sequence of a novel member of the genus Caulimovirus isolated from Dregea volubilis.

A novel caulimovirus was identified from diseased leaves of Dregea volubilis exhibiting yellowing and vein-associated chlorosis in Yuanjiang County, Yunnan Province, China. The virus was tentatively named Dregea volubilis caulimovirus 1 (DVCaV1). The complete genome sequence of DVCaV1, determined by de novo assembly of high-throughput sequencing data, comprises 8,160 bp of circular double-stranded DNA containing two intergenic regions and seven open reading frames (ORFs). These ORFs encode (in order) a movement protein (MP), an aphid transmission factor (ATF), a virion-associated protein (VAP), a coat protein (CP), a polymerase polyprotein (Pol, containing protease, reverse transcriptase, and RNase H domains), a transactivator/viroplasmin (TAV) protein, and a hypothetical protein of unknown function. Sequence comparisons revealed the highest nucleotide similarity with strawberry vein banding virus (SVBV; NC_001725). Phylogenetic analysis confirmed DVCaV1 as a member of the genus Caulimovirus, with SVBV as its closest known relative. According to current ICTV species demarcation criteria for the genus Caulimovirus (host range and > 20% nucleotide sequence divergence in the polymerase region), DVCaV1 represents a novel species. This is, to our knowledge, the first report of a caulimovirus detected in naturally symptomatic Dregea volubilis.

Genome, Viral↗
PubMed1993

The putative zinc finger of a caulimovirus is essential for infectivity but does not influence gene expression.

Plant pararetroviruses, such as caulimoviruses, and animal retroviruses have in common the presence of a highly conserved arrangement of cysteines and a histidine in the precursor of the capsid protein. The composition of these amino acids resembles a zinc finger element, a structure that is common to a class of eukaryotic proteins that regulate gene expression. The role of the putative zinc finger in the life-cycle of caulimoviruses was investigated by introducing specific mutations in the coat protein coding region of a cloned and infectious form of figwort mosaic virus, a caulimovirus. This mutated viral genome, which no longer encoded the conserved cysteine and histidine residues, was not infectious in plants. Transient expression assays in protoplasts showed that expression of a reporter gene inserted at different places in the genome was not detectably influenced by the coat protein or its putative zinc finger. It appears that the zinc finger-like element of caulimoviruses is not involved in the regulation of gene expression. These observations support a model which predicts a function of the zinc finger in specific recognition and packaging of viral RNA into virions prior to reverse transcription.

Amino Acid Sequence↗
PubMed1998

A common 40 amino acid motif in eukaryotic RNases H1 and caulimovirus ORF VI proteins binds to duplex RNAs.

Eukaryotic RNases H from Saccharomyces cerevisiae , Schizosaccharomyces pombe and Crithidia fasciculata , unlike the related Escherichia coli RNase HI, contain a non-RNase H domain with a common motif. Previously we showed that S.cerevisiae RNase H1 binds to duplex RNAs (either RNA-DNA hybrids or double-stranded RNA) through a region related to the double-stranded RNA binding motif. A very similar amino acid sequence is present in caulimovirus ORF VI proteins. The hallmark of the RNase H/caulimovirus nucleic acid binding motif is a stretch of 40 amino acids with 11 highly conserved residues, seven of which are aromatic. Point mutations, insertions and deletions indicated that integrity of the motif is important for binding. However, additional amino acids are required because a minimal peptide containing the motif was disordered in solution and failed to bind to duplex RNAs, whereas a longer protein bound well. Schizosaccharomyces pombe RNase H1 also bound to duplex RNAs, as did proteins in which the S.cerevisiae RNase H1 binding motif was replaced by either the C.fasciculata or by the cauliflower mosaic virus ORF VI sequence. The similarity between the RNase H and the caulimovirus domain suggest a common interaction with duplex RNAs of these two different groups of proteins.

Amino Acid Sequence↗
PubMed1995

Gene I mutants of peanut chlorotic streak virus, a caulimovirus, replicate in plants but do not move from cell to cell.

Gene I of peanut chlorotic streak virus (PCISV), a caulimovirus, is homologous to gene I of other caulimoviruses and may encode a protein for virus movement. To evaluate the function of gene I, several mutations were created in this gene of an infectious, partially redundant clone of PCISV. Constructs with an in-frame deletion and a single amino acid substitution in gene I were not infectious. To test for replication of these mutants in primarily infected cells, an immunosorbent PCR technique was devised. Virus particles formed by mutants in plants were recovered by binding to antivirus antibodies on a solid matrix and DNase treated to discriminate against residual inoculum, and DNA of trapped virions was subjected to PCR amplification. Gene I mutants were shown to direct formation of encapsidated DNA as revealed by a PCR product. Control gene V mutants (reverse transcriptase essential for replication) did not yield a PCR product. Quantitative PCR allowed estimation of the proportion of cells initially infected by gene I mutants and the amount of extractable virus per cell. It is concluded that PCISV gene I encodes a movement protein and that the immunoselection-PCR technique is useful in studying subliminal virus infection in plants.

Amino Acid Sequence↗
PubMed1991

The regions of sequence variation in caulimovirus gene VI.

The sequence of gene VI from figwort mosaic virus (FMV) clone x4 was determined and compared with that previously published for FMV clone DxS. Both clones originated from the same virus isolation, but the virus used to clone DxS was propagated extensively in a host of a different family prior to cloning whereas that used to clone x4 was not. Differences in the amino acid sequence inferred from the DNA sequences occurred in two clusters. An N-terminal conserved region preceded two regions of variation separated by a central conserved region. Variation in cauliflower mosaic virus (CaMV) gene VI sequences, all of which were derived from virus isolates from hosts from one host family, was similar to that seen in the FMV comparison, though the extent of variation was less. Alignment of gene VI domains from FMV and CaMV revealed regions of amino acid sequence identical in both viruses within the conserved regions. The similarity in the pattern of conserved and variable domains of these two viruses suggests common host-interactive functions in caulimovirus gene VI homologues, and possibly an analogy between caulimoviruses and certain animal viruses in the influence of the host on sequence variability of viral genes.

Amino Acid Sequence↗
PubMed1992

The full-length transcript of a caulimovirus is a polycistronic mRNA whose genes are trans activated by the product of gene VI.

Gene expression of figwort mosaic virus (FMV), a caulimovirus, was investigated by electroporation of Nicotiana edwardsonii cell suspension protoplasts with cloned viral constructs in which a reporter gene was inserted at various positions on the genome. The results showed that the genome of FMV contains two promoters; one is used for the production of a full-length RNA and another initiates synthesis of a separate monocistronic RNA for gene VI. Evidence is provided that the full-length transcript, the probable template for reverse transcription, can serve as a polycistronic mRNA for translation of genes I through V and perhaps also gene VI. Expression of all the genes on the polycistronic mRNA is trans activated by the gene VI protein. Reporter gene expression appears most efficient when its start codon is in close proximity to the stop codon of the preceding gene, as for the native genes of caulimoviruses. We propose that the gene VI product enables expression of the polycistronic mRNA by promoting reinitiation of ribosomes to give translational coupling of individual genes.

Cells, Cultured↗
PubMed1986

The molecular biology of caulimoviruses.

The molecular biology of the caulimoviruses has already inspired quite a few review articles: this review is limited to a general description of the type-member of this group, namely cauliflower mosaic virus. Details are presented of major results obtained on the organization and function at the molecular level of caulimoviruses and cauliflower mosaic virus in particular.

Base Sequence↗
PubMed1998

Isolation and expression analysis of peanut chlorotic streak caulimovirus (PClSV) full-length transcript (FLt) promoter in transgenic plants.

A promoter fragment from peanut chlorotic streak caulimovirus (PClSV) full-length transcript (FLt) was identified and later modified to have duplicated enhancer domain. The FLt promoter with its single or double enhancer domains, fused with the GUS reporter gene to form chimeric gene constructs, showed a high level of expression of these genes in cells and transgenic plants. The FLt promoter with its double enhancer domain gives an average threefold greater expression of genes compared to the FLt promoter with its single enhancer domain in transgenic plants. In young seedlings the expression was in the order root > leaf > stem. The histochemical GUS assay in young seedlings showed more activity in root tips and leaf midribs, veins, and other vascular tissues. The expression from the PClSV FLt promoter was compared with that from the figwort mosaic virus promoter in transgenic plants. These constitutive promoters were comparable in respect to GUS expression level.

Caulimovirus↗
PubMed1998

Gene expression regulated by gene VI of caulimovirus: transactivation of downstream genes of transcripts by gene VI of peanut chlorotic streak virus in transgenic tobacco.

Here we document that the gene VI product of peanut chlorotic streak virus (PClSV), a newly characterized member of the group, transactivates the translation of dicistronic transcripts. Dicistronic expression units have been analyzed both in protoplast transient expression experiments and in transgenic tobacco plants. Transgenic plants containing a dicistronic transcription unit (PClSV-gene VII-GUS) under the control of PClSV full-length transcript promoter with its long leader sequence show a relatively high abundance of the expected transcript but very little, or no, GUS activity. However, high GUS activity is found when gene VI protein is then provided by subsequent infection with PClSV. The efficient translation of polycistronic mRNAs mediated by gene VI of caulimovirus has potential value in product engineering of plants.

Base Sequence↗
PubMed1998

Strawberry vein banding virus--definitive member of the genus Caulimovirus.

The complete DNA sequence (7876 nucleotides) of strawberry vein banding virus (SVBV) has been determined. Seven open reading frames are detected that potentially code for proteins of calculated weight 37.8; 18.3; 16.6; 56.0; 81.1; 59.0 and 12.6 kDa, respectively. Their position on the viral genome is the same as that of the corresponding proteins on the cauliflower mosaic virus (CaMV) genome. Phylogenetic analysis based on the amino acid sequence of this protein shows a closer relationship of SVBV with CaMV, figwort mosaic virus and carnation etched ring virus than with other caulimoviruses.

Base Sequence↗
PubMed1994

Eukaryotic RNAse H shares a conserved domain with caulimovirus proteins that facilitate translation of polycistronic RNA.

RNAse H (RNH1 protein) from the trypanosomatid Crithidia fasciculata has a functionally uncharacterized N-terminal domain dispensable for the RNAse H activity. Using computer methods for database search and multiple alignment, we show that the N-terminal domains of RNH1 and its homologue encoded by a cDNA from chicken lens are related to the conserved domain in caulimovirus ORF VI product that facilitates translation of polycistronic virus RNA in plant cells. We hypothesize that the N-terminal domain of eukaryotic RNAse H performs an as yet uncharacterized regulatory function, possibly in mRNA translation or turnover.

Amino Acid Sequence↗
PubMed1996

Solid-phase synthesis, metal binding and folding properties of caulimovirus-related 'zinc finger'.

A 17-residue peptide containing the caulimovirus-related "zinc finger' was prepared by solid-phase peptide synthesis. Fluorescence measurements showed that the tryptophan quantum yield was Zn(2+)-dependent, allowing a 1:1 a stoichiometry for the complex to be determined. The structure of the peptide was characterized using circular dichroic spectroscopy, which indicates that the peptide exhibits a random coiled conformation in the absence of zinc but appears to form an ordered structure in the presence of zinc.

Amino Acid Sequence↗
PubMed1997

Multiple widely spaced elements determine the efficiency with which a distal cistron is expressed from the polycistronic pregenomic RNA of figwort mosaic caulimovirus.

The polycistronic expression mechanism of the plant pararetrovirus figwort mosaic caulimovirus (FMV) depends upon cis-acting elements present in its pregenomic RNA and a trans-acting protein (P6) which is expressed from a monocistronic subgenomic RNA. Using transient expression of FMV-derived polycistronic reporter constructs in Nicotiana edwardsonii cell suspension protoplasts, we further analyzed the cis-acting elements involved in polycistronic expression. A cis-acting element located within the first 74 nucleotides of the 7,954-nucleotide pregenomic RNA appears to be essential for P6 to transactivate expression of an internal cistron. Expression of this internal cistron, in the presence of P6, is greatly enhanced by the combined presence of two cis-acting elements located at the 3' end of the polycistronic RNA. Surprisingly, deletion of the most upstream of these two 3' cis-acting elements exposed a negative-acting element located internally on the polycistronic RNA, at the 3' end of open reading frame I. The action of both this negative-acting internal element and the positive-acting 3' elements is more pronounced when the large 5' untranslated leader region is present. This indicates that the 5' untranslated leader region is central to regulation of the FMV gene expression mechanism. Although a limited set of elements suffices to direct polycistronic expression in this eukaryotic system, a complex interplay between elements is involved in the spatial regulation of the genes present on the pregenomic RNA of FMV.

Caulimovirus↗
PubMed1991

Characterization of the genome of rice tungro bacilliform virus: comparison with Commelina yellow mottle virus and caulimoviruses.

Rice tungro disease is caused by an infection of two different viruses, rice tungro spherical virus (a (+) sense RNA virus) and rice tungro bacilliform virus (RTBV) with a genome of circular double-stranded DNA. The genome of an RTBV isolate from the Philippines was cloned, sequenced, and found to be 8000 bp in length. It contains four open reading frames (ORFs) on a single strand, with ORF 1 having an internal termination codon (TAA). The 5' and 3' ends of a polyadenylated viral RNA transcript, of genome length, were mapped by primer extension and cDNA sequence analysis, respectively. The transcript is terminally redundant by 265-268 nucleotides. Purified virus particles contain two major proteins with molecular masses of 37 and 33 kDa, although only the 37-kDa protein was detected in the infected rice tissues. The N-terminal amino acid sequence of the 33-kDa protein was determined and its coding region was identified on the RTBV genome. The identity of the coat protein gene was further confirmed by expressing a region of the genome in Escherichia coli, the products of which reacted with anti-RTBV antibody. The unusually long ORF 3 of RTBV is predicted to encode a polyprotein of 194.1 kDa that includes: the coat protein(s), viral proteinase, reverse transcriptase, and ribonuclease H. The sections of the polyprotein show varying degrees of similarity to the counterparts of Commelina yellow mottle virus (a member of the proposed badnavirus group) and caulimoviruses. The functions of the other three ORFs are unknown.

Amino Acid Sequence↗
PubMed1991

A naturally occurring deletion mutant of figwort mosaic virus (caulimovirus) is generated by RNA splicing.

A naturally occurring deletion mutant is observed in plants infected with figwort mosaic virus (FMV), a caulimovirus. The encapsidated mutant genome is formed spontaneously in association with two different strains of FMV in four host plant species. The mutant also appears when cloned wild-type viral DNA is used as the inoculum. The deletion mutant alone is not infectious and it appears unable to replicate after its formation, even in the presence of wild-type virus. The gene for chloramphenicol acetyltransferase was inserted at different positions in the deletion mutant genome, and subsequent transient assays showed that gene expression of the mutant occurs despite the deletion. Sequence analyses of the mutant genome revealed a deletion of 1237-bp segment encompassing a major portion of the coat protein gene and the 5' end of the downstream reverse transcriptase gene. This deletion is associated with consensus signals for RNA splicing including the conserved 5' and 3' splice sites plus surrounding sequences, putative branch point(s) for lariat formation, and an extremely high adenosine content (41%) of the removed fragment. This suggests that splicing of the FMV full-length transcript has occurred prior to reverse transcription and this accounts for the presence and accumulation of encapsidated DNAs with the same deletion.

Base Sequence↗
PubMed1992

Regulation of caulimovirus gene expression and the involvement of cis-acting elements on both viral transcripts.

In a further analysis of gene regulation of figwort mosaic virus (FMV), a caulimovirus, we studied transient gene expression with modified viral genomes in Nicotiana edwardsonii cell suspension protoplasts. The results demonstrated that the presence of the promoter for the full-length RNA interferes with expression from the separate downstream promoter for gene VI. In addition, expression of gene VI was inhibited by cis-acting sequences within gene VI itself. Both inhibitory effects could be partially relieved by coelectroporation with a plasmid that produces gene VI protein, demonstrating that expression of gene VI is transactivated by its own product. Subsequent expression studies with partially redundant FMV plasmids containing a reporter gene in frame with gene IV showed that efficient transactivation of CAT expression relies on a cis-acting element inside the downstream gene VI. Insertions of a transcriptional terminator upstream of the cis-acting element for premature termination of transcription showed that the cis-acting region is not a DNA element but is active only as a feature of the RNA transcript. We conclude that the cis-acting element, together with the transacting gene VI product, enhances expression of all major genes, including gene VI, from the polycistronic mRNA and the separate mRNA for gene VI.

Enhancer Elements, Genetic↗
PubMed1989

Gene VI of figwort mosaic virus (caulimovirus group) functions in posttranscriptional expression of genes on the full-length RNA transcript.

Experimental evidence for a molecular function for gene VI of the caulimoviruses is presented. Based on experiments with the figwort mosaic virus (FMV), it appears that gene VI has a role in the posttranscriptional expression of the closely packed genes (VII and I-V), which appear on the larger, full-length RNA transcript of this virus. Gene VI with its flanking 5'/3' expression signals included as a separate plasmid during electroporation of DNA into protoplasts of Nicotiana edwardsonii shows an unusual type of transactivation of a chloramphenicol acetyltransferase (CAT) gene fused at its 5' end to a small open reading frame (gene VII) of the long 5' leader of the full-length RNA transcript of the FMV genome. The level of activity of the CAT gene is increased up to 20-fold over the activity of control plasmids when gene VI is included in the electroporation mixture. Mutagenesis of the coding portions of gene VI of pGS1 RVI, a transactivating plasmid used in the electroporation experiments, demonstrated that it was probably the polypeptide product of gene VI that was responsible for the transactivating effect. Experiments with various portions of the 5' leader of the large, full-length RNA of FMV showed that the coding region of gene VII is necessary for the transactivation event. Clones of cauliflower mosaic virus (CaMV) or FMV with intact gene VI were found to reciprocally transactivate gene VII-CAT fusions (FMV) or gene I-CAT fusions (CaMV) located downstream of the 5' leader sequences of either viral genome.

Base Sequence↗