Genetic Determinants of Symptoms on Viral DNA Satellites

Abstract

Begomovirus-DNA-␤ disease complexes induce different symptom phenotypes in their hosts. To investigate the genetic determinants of the phenotypic differences, Nicotiana spp. and tomato plants were inoculated with infectious clones of Tobacco curly shoot virus (TbCSV)/TbCSV DNA-␤ (TbCSB) and Tomato yellow leaf curl China virus (TYLCCNV)/TYLCCNV DNA-␤ (TYLCCNB) pseudorecombinants and showed that TYLCCNB induced characteristic vein-thickening and enation symptoms, while TbCSB only slightly exacerbated the leaf-curling symptoms, regardless of the helper virus being used. The roles of DNA-␤-encoded ␤C1 and a 430-nucleotide fragment containing the A-rich region and the putative ␤C1 promoter region of the ␤C1 gene (referred to as AP) in symptom development were further investigated by constructing hybrid satellites in which the ␤C1 coding region or AP was exchanged between the two satellite molecules. A TYLCCNB hybrid with TbCSB ␤C1 lost the ability to elicit the vein-thickening and enation phenotypes. TbCSB hybrids containing the TYLCCNB ␤C1 or AP fragment failed to induce the characteristic vein thickening and enations. A TYLCCNB hybrid having the TbCSB AP fragment produced the enations, but the number of enations was less and their sizes were reduced. Differently from the phloem-specific pattern of the TYLCCNB promoter, a full-length fragment upstream of the TbCSB ␤C1 gene confers a constitutive ␤-glucuronidase expression pattern in transgenic tobacco plants. The above results indicate that the DNA-␤-encoded ␤C1 protein is the symptom determinant, but the promoter of the ␤C1 gene has influence on symptom production.Geminiviruses are small plant viruses with circular singlestranded DNA (ssDNA) genomes that are encapsidated in unique twinned (geminate) particles. Members of the genus Begomovirus are transmitted by whiteflies (Bemisia tabaci) and infect dicotyledonous plants (42). Begomoviruses have either one or two circular ssDNA genomic components (DNA-A and DNA-B). The DNA-A component is capable of autonomous replication and encapsidation, whereas the DNA-B component encodes two proteins (BC1 and BV1) involved in movement (14). Recently, some monopartite begomoviruses have been found in association with a novel satellite DNA molecule, referred to as DNA-␤ and now known as a betasatellite (2,5,20,22,38,45). DNA-␤ is approximately half the size of the viral genomic DNA, and apart from a nonanucleotide sequence (TA ATATTAC), it has little sequence identity with viral genomic DNA. DNA-␤ depends on the helper virus for replication and encapsidation and, in turn, is required for the induction of bona fide disease symptoms. DNA-␤ bears a ␤C1 open reading frame (ORF) on the complementary-sense strand, which is conserved among distinct betasatellites in terms of position and size. Mutational analyses and constitutive expression have shown that ␤C1 is a strong pathogenicity/symptom determinant (7, 34, 39).Begomovirus-DNA-␤ disease complexes are associated with a wide range of plant species and induce different sets of sympto...

Suppression of RNA Silencing by a Plant DNA Virus Satellite Requires a Host Calmodulin-Like Protein to Repress RDR6 Expression

“…While able to infect their hosts alone, these monopartite begomoviruses are often incapable of inducing typical symptoms, and require betasatellites for accumulation to high titers and elicitation of disease symptoms [39], [40]. The betasatellite contains a single open reading frame (βC1) that encodes a symptom determinant [41]–[44].…”

A Novel DNA Motif Contributes to Selective Replication of a Geminivirus-Associated Betasatellite by a Helper Virus-Encoded Replication-Related Protein

“…Infectious clones of the hybrid betasatellites, pBinPLUS-Y10␤-35␤C1, pBinPLUS-Y35␤-10␤C1, pBinPLUS-Y10␤-35AP, and pBinPLUS-Y35␤-10AP (referred to here as Y10␤-35␤C1, Y35␤-10␤C1, Y10␤-35AP, and Y35␤-10AP, respectively), were also described previously (29). Other hybrid betasatellite constructs were produced as previously described (29). Briefly, a splicing overlap extension PCR (SOE-PCR) strategy was employed to precisely exchange the LCR (left side of the SCR, a 260-nt fragment upstream of the primer pair ␤01 and ␤02 located positions), the RCR (right side of the SCR, a 270-nt fragment downstream of the positions of the primer pair ␤01 and ␤02), and the RBM (Rep-binding motif) between Y10␤ and Y35␤ or to delete the RBM of the betasatellite (Fig.…”

“…The infectious clones of TYLCCNV isolate Y10 (pBinPLUS-Y10-1.7A) and its betasatellite TYLCCNB (pBinPLUS-Y10-2␤), TbCSV isolate Y35 (pBinPLUS-Y35-1.7A) and its betasatellite TbCSB (pBinPLUS-Y35-2␤), and tomato leaf curl China virus isolate G18 (ToLCCNV, pBinPLUS-G18-1.7A) and its betasatellite ToLCCNB (pBinPLUS-G18-2␤) (referred to here as Y10 and Y10␤, Y35 and Y35␤, and G18 and G18␤, respectively) were constructed previously (7,27,28). Infectious clones of the hybrid betasatellites, pBinPLUS-Y10␤-35␤C1, pBinPLUS-Y35␤-10␤C1, pBinPLUS-Y10␤-35AP, and pBinPLUS-Y35␤-10AP (referred to here as Y10␤-35␤C1, Y35␤-10␤C1, Y10␤-35AP, and Y35␤-10AP, respectively), were also described previously (29). Other hybrid betasatellite constructs were produced as previously described (29).…”

Viral Metagenomics: Analysis of Begomoviruses by Illumina High-Throughput Sequencing

“…These phenomena are in part made possible by the suppressor activity of the betasatellite-encoded protein, β C 1 that silences the plant host response to viral infection [8]. Further betasatellites are encapsidated into virions for transmission from plant to plant, along with the helper virus, by the whitefly vector Bemisia tabaci (Gennadius) sibling species group [9,10].…”