The oligomeric Rep protein of Mungbean yellow mosaic India virus (MYMIV) is a likely replicative helicase

Geminiviruses replicate by rolling circle mode of replication (RCR) and the viral Rep protein initiates RCR by the site-specific nicking at a conserved nonamer (TAATATT downward arrow AC) sequence. The mechanism of subsequent steps of the replication process, e.g. helicase activity to drive fork-elongation, etc. has largely remained obscure. Here we show that Rep of a geminivirus, namely, Mungbean yellow mosaic India virus (MYMIV), acts as a replicative helicase. The Rep-helicase, requiring > or =6 nt space for its efficient activity, translocates in the 3'-->5' direction, and the presence of forked junction in the substrate does not influence the activity to any great extent. Rep forms a large oligomeric complex and the helicase activity is dependent on the oligomeric conformation ( approximately 24mer). The role of Rep as a replicative helicase has been demonstrated through ex vivo studies in Saccharomyces cerevisiae and in planta analyses in Nicotiana tabacum. We also establish that such helicase activity is not confined to the MYMIV system alone, but is also true with at least two other begomoviruses, viz., Mungbean yellow mosaic virus (MYMV) and Indian cassava mosaic virus (ICMV).     

Characterization of Mungbean yellow mosaic India virus genome with a recombinant DNA-B in Southern Peninsular India.

Molecular Biology Reports - Mungbean yellow mosaic India virus (MYMIV) is a representative of the genus begomovirus/Begomoviridae, which is prevalent in the northern part of Indian subcontinent...     

Diagnosis of Symptomless Yellow mosaic begomovirus Infection in Pigeonpea by Using Cloned Mungbean yellow mosaic India virus as Probe

One isolate of Mungbean yellow mosaic India virus (MYMIV) of mungbean plants from Sri Ganganagar, Rajasthan, designated as MYMIV-Mg was isolated and DNA-A and DNA-B, the two full length bipartite genomic components of this virus, were cloned. The [α-³²P] labeled diagnostic probes specific to these cloned DNA-A and -B of MYMIV-Mg were used to detect the virus infection in infected plants by nucleic acid spot hybridization (NASH) test. The NASH tests detected the MYMIV infection and concentration of viral titre in susceptible, moderately susceptible, resistant and symptomless genotypes of pigeonpea (Cajanus cajan) plants. Fourteen genotypes of pigeonpea were tested against five naturally occurring MYMIV variants viz.,.MYMIV Bg, -MgD, -MoL, -Mg and -Pp1 through viruliferous whitefly (Bemisia tabaci) transmission in greenhouse condition. Disease incidence and severity of MYMIV in different pigeonpea genotypes varied with the variants of MYMIV. Many genotypes of pigeonpea did not produce visible yellow mosaic symptoms after inoculation with MYMIV variants MYMIV-Bg, -MbD and -MoL, although, majority of the symptomless genotypes were found to be infected by MYMIV, as viral DNA was detected by NASH test.     

The 32 kDa subunit of replication protein A (RPA) participates in the DNA replication of Mung bean yellow mosaic India virus (MYMIV) by interacting with the viral Rep protein

Mung bean yellow mosaic India virus (MYMIV) is a member of genus begomoviridae and its genome comprises of bipartite (two components, namely DNA-A and DNA-B), single-stranded, circular DNA of about 2.7 kb. During rolling circle replication (RCR) of the DNA, the stability of the genome and maintenance of the stem–loop structure of the replication origin is crucial. Hence the role of host single-stranded DNA-binding protein, Replication protein A (RPA), in the RCR of MYMIV was examined. Two RPA subunits, namely the RPA70 kDa and RPA32 kDa, were isolated from pea and their roles were validated in a yeast system in which MYMIV DNA replication has been modelled. Here, we present evidences that only the RPA32 kDa subunit directly interacted with the carboxy terminus of MYMIV-Rep both in vitro as well as in yeast two-hybrid system. RPA32 modulated the functions of Rep by enhancing its ATPase and down regulating its nicking and closing activities. The possible role of these modulations in the context of viral DNA replication has been discussed. Finally, we showed the positive involvement of RPA32 in transient replication of the plasmid DNA bearing MYMIV replication origin using an in planta based assay.     

Antisense RNA approach targeting Rep gene of Mungbean yellow mosaic India virus to develop resistance in soybean

The Yellow mosaic disease is caused by Mungbean yellow mosaic India virus (MYMIV) and Mungbean yellow mosaic virus (MYMV) belonging to the genus Begomovirus of the family Geminiviridae. Yellow mosaic disease (YMD) is a major constraint to the production of soybean in South-East Asia. In India, yield losses of 10–88% had been reported due to YMD of soybean. An effort has been made to generate resistant soybean plants, by a construct targeting replication initiation protein (Rep) gene sequences of MYMIV. A construct containing the sequences of Rep gene (566?bp) in antisense orientation was used to transform cotyledonary node explants of three soybean cultivars (JS 335, JS 95-60 and NRC 37). Transformation efficiencies of 0.2, 0.21 and 0.24% were obtained with three soybean cultivars, JS 335, JS 95-60 and NRC 37, respectively. The presence of transgene in T₁ plants was confirmed by polymerase chain reaction (PCR) and sequence analysis. The level of resistance was observed by challenge inoculation with the virus in T₁ lines. The inheritance of transgene showed classical Mendelian pattern in six transgenic lines.     

Characterisation of pumpkin yellow vein mosaic virus from India

Yellow vein mosaic disease symptoms occur frequently in pumpkin in India. Diseased plants show vein yellowing, which sometimes coalesces to form chlorotic patches. Infected plants are stunted and flowers drop prematurely, greatly reducing yields. Diseased plants are infected by a begomovirus, designated pumpkin yellow vein mosaic virus (PYVMV), which is transmitted readily and in a persistent manner by the whitefly, Bemisia tabaci. Transmission of PYVMV requires minimum acquisition and inoculation access periods of 30 min and 10 min, respectively. The minimum latent period in the insect is 6 h and the virus persists in the vector for at least 8 days. PYVMV has a narrow host range consisting of a small number of cucurbit species and some tobacco cultivars. It was detected serologically in diseased plants and in viruliferous B. tabaci using polyclonal antibodies in a double‐antibody sandwich enzyme‐linked immunosorbent assay. Reactions with monoclonal antibodies in a triple‐antibody sandwich ELISA showed that PYVMV has an epitope profile distinct from those of other begomoviruses from the Indian sub‐continent. Polymerase chain reaction amplified fragments from the putative viral coat and movement protein genes. Based on comparative phylogeny of complete coat protein gene sequences, PYVMV was most similar to the bipartite Tomato leaf curl New Delhi virus from India and appears to be a new strain of this virus.     

Coat protein polymerization of alfalfa mosaic virus strain VRU

The association behaviour of the coat protein of alfalfa mosaic virus strain VRU was studied by sedimentation analysis and electron microscopy. The results of this study were compared with the data obtained from similar studies with the coat protein of strain 425 (Driedonks et al., 1977). In the depolymerized state VRU protein is likely a dinier of the 24,050 molecular weight polypeptide chain. The main association product is a tubular structure with a diameter of about 180 Å. The optimum conditions for the reaction were polyphosphate-containing buffer at pH 6·5. Optical diffraction analysis of negatively stained specimens revealed a helical arrangement of the protein subunits in these assemblies. The same type of reaction product was found when the association reaction was carried out in the presence of polynucleotides. The length of the VRU particles is abnormally long compared to other alfalfa mosaic virus strains. This phenomenon can be ascribed to the tendency of the protein to polymerize into tubular rather than spherical particles.     

The DNA-1 like alphasatellites in association with the bipartite Mungbean yellow mosaic virus in natural infections

Yellow mosaic disease is the major limitation in the production of grain legumes in India. This disease is caused by bipartite begomovirus, Mungbean yellow mosaic virus. In addition to the bipartite genomic components, the yellow mosaic disease affected urdbean plants which contain satellite like DNA-1 component called as alphasatellites. The present study has been attempted to characterise the alphasatellites associated with Mungbean yellow mosaic virus. Nucleotide sequence analysis of alphasatellites showed 98% identity with Vernonia yellow vein Fijian alphasatellite, VYVFA (JF733780). Since the sequence identity is more than 98%, the threshold value for demarcation of alphasatellites species, the alphasatellites of the present study are named as Vernonia yellow vein Fijian alphasatellite. Comparison with other, alphasatellites shared 51–55% identity with alphasatellites associated with monopartite begomovirus and it shared only 41–42% identity with an unusual alphasatellites, DNA-2. This is the first report on characterisation of alphasatellites associated with Mungbean yellow mosaic virus.     

Nucleotide sequence of turnip yellow mosaic virus coat protein mRNA

The primary structure of the coat protein messenger RNA of turnip yellow mosaic virus is presented. This sequence is the first complete nucleotide sequence of the coat protein messenger of a plant virus to be reported. The coding region, consisting of 567 nucleotides, is flanked by a 5′ noncoding region of 19 nucleotides (not including the initiation codon and the cap structure) and by a 3′ noncoding region of 109 nucleotides (including the termination signal). The coat protein mRNA has a base composition identical to that of the genome RNA with, in particular, the same high content in cytosine (38%). The codons that govern the incorporation of amino acids into the coat protein are nonrandomly utilized: >50% of the time the third base of the codons used is a cytosine. This pattern of codon preference is particularly marked for Leu, lie, Val, Thr and Cys.     

Identification of Mungbean yellow mosaic Indian virus Associated with Tomato Leaf Curl Betasatellite Infecting Phaseolus vulgaris in Oman

Kidney bean (Phaseolus vulgaris) plants exhibiting foliar yellow mosaic symptoms and some leaf crumpling were identified in the Al Batinah region of Oman. Rolling circle amplification and polymerase chain reaction identified a bipartite begomovirus (family Geminiviridae) and a betasatellite in association with the symptomatic plants. Analysis of full‐length sequences showed the virus to be Mungbean yellow mosaic Indian virus (MYMIV) and the betasatellite Tomato leaf curl betasatellite (ToLCB). This is the first identification of a legume‐adapted begomovirus in Oman and the first identification of MYMIV in association with the betasatellite ToLCB. The isolate of MYMIV from Oman shows the greatest levels of sequence identity to isolates occurring in South Asia and South‐East Asia, suggesting that the virus has only recently been introduced. The significance of these findings is discussed.     

Reactions of some cucurbitaceous species Tozucchini yellow mosaic virus (ZYMV)

Zucchini yellow mosaic virus (ZYMV) is a widespread serious pathogen of cucurbitaceous plants. ZYMV was first detected in Hungary in 1995. Since then it has become one of the most dangerous viruses of the Cucurbitaceae family causing serious epidemics. The virus has many hosts, which - particularly perennial ones - may play important role as virus reservoirs and infection sources in virus epidemiology. On the other hand wild weed species maybe sources of resistance to viruses. Our research was carried out on a total of 15 wild species from 8 genera (Cucumis, Cucurbita, Cyclanthera, Ecballium Momordica, Lagenaria, Zehneria, Bryonia). Test plants were mechanically inoculated with ZYMV. Local and systemic symptoms were determined and 5 weeks after inoculation DAS-ELISA tests were also carried out. Symptomless plants were reinoculated to Cucumis sativus cv. Accordia test plants. On the basis of the results we determined the percentages of infections and so we classified the test-plants into sensitive and resistance categories. On the basis of the results new host plants of ZYMV are the followings: Bryonia dioica, Cyclanthera pedata, Ecballium elaterium, Momordica balsamina, Momordica rostrata, and Zehneria scabra. Among them Momordica balsamina and Ecballium elaterium showed latent to ZYMV. Bryonia alba and Zehneria indica are especially remarkable, because they proved resistant to ZYMV on the basis of symptomatology and serology. Our results might have significant role in the field of research of host range, virus resistance and virus differentiation.     

Differential responses of Phaseolus vulgaris cultivars following mungbean yellow mosaic India virus infection

Physiology and Molecular Biology of Plants - Phaseolus vulgaris, commonly known as French bean is a vital leguminous crop worldwide and India stood 1st rank in dry bean and 4th rank in green bean...     

First Report of a Croton yellow vein mosaic virus (CYVMV) Associated with Tomato Leaf Curl Disease in India

Leaf curl disease symptoms were observed in tomato crop grown in a tomato field at Matera district of Bahraich, India, in March 2013 with an 85% disease incidence. The infected plants exhibited leaf curl symptoms accompanied with puckering, vein swelling and stunting of the whole plant. PCR carried out with begomovirus coat protein gene and DNA beta‐specific primer sets resulted in positive amplification of ~775 bp and 1.35 kbp, respectively, with all symptom‐bearing plant samples. BLASTn and phylogenetic analyses of CP gene sequences showed highest and close relationship with Croton yellow vein mosaic virus (CYVMV) isolates, while the phylogenetic study of betasatellite sequence showed distinct relationships with other begomovirus associated betasatellites reported from India and abroad. This is a first report of a CYVMV associated with tomato leaf curl disease in India.     

Sequencing of Coat Protein Gene of an isolate of Cucumber mosaic virus Infecting Black Pepper (Piper nigrum L) in India

The coat protein gene of an isolate of Cucumber mosaic virus (CMV) associated with stunted disease affecting black pepper plant was cloned and sequenced. The coat protein gene comprised of 657 nucleotides encoding a protein of 218 amino acids. Sequence analysis showed that the gene was most closely related to CMV infecting Egyptian henbane plant in India, the member of subgroup I of CMV. The amino acid sequence identity with members of subgroup I was 92-99% while that with members of subgroup II was 77–79%. On the basis of sequence homology, it is concluded that CMV infecting black pepper in India is a strain of CMV belonging to subgroup I.