Cauliflower Mosaic Virus Aphid Transmission Research Articles

The N-terminus of the cauliflower mosaic virus aphid transmission protein P2 is involved in transmission body formation and microtubule interaction

Cauliflower mosaic virus (CaMV) is transmitted by aphids using the non-circulative transmission mode: when the insects feed on infected leaves, virus particles from infected cells attach rapidly to their stylets and are transmitted to a new host when the aphids change plants. Mandatory for CaMV transmission, the viral helper protein P2 mediates as a molecular linker binding of the virus particles to the aphid stylets. P2 is available in infected plant cells in a viral inclusion that is specialized for transmission and named the transmission body (TB). When puncturing an infected leaf cell, the aphid triggers an ultra-rapid viral response, necessary for virus acquisition and called transmission activation: The TB disrupts and P2 is redistributed onto cortical microtubules, together with virus particles that are simultaneously set free from virus factories and join P2 on the microtubules to form the so-called mixed networks (MNs). The MNs are the predominant structure from which CaMV is acquired by aphids. However, the P2 domains involved in microtubule interaction are not known. To identify P2 regions involved in its functions, we generated a set of P2 mutants by alanine scanning and analyzed them in the viral context for their capacity to form a TB, to interact with microtubules and to transmit CaMV. Our results show that contrary to the previously characterized P2-P2 and P2-virion binding sites in its C-terminus, the microtubule binding site is contained in the N-terminal half of P2. Further, this region is important for TB formation since some P2 mutant proteins did not accumulate in TBs but were retained in the viral factories where P2 is translated. Taken together, the N-terminus of P2 is not only involved in vector interaction as previously reported, but also in interaction with microtubules and in formation of TBs.

Molecular dynamics investigations for the prediction of molecular interaction of cauliflower mosaic virus transmission helper component protein complex with Myzus persicae stylet's cuticular protein and its docking studies with annosquamosin-A encapsulated in nano-porous Silica.

Large numbers of bioactive natural products from plant species such as alkaloids, phenolics, terpenoids etc. are remaining unexplored for their potential as plant protective agents as inhibitors for viral and other pathogenic infections of plant. Myzus aphids are important plant pests and vectors for several plant viruses. Cauliflower mosaic virus (CaMV) belongs to the plant virus family Caulimoviridae which is transmitted "non-circulative" from plant to plant through an interaction with aphid insect vectors. This viral transmission process most likely involves a protein-protein binding interaction between aphid stylet receptor cuticular protein and viral proteins namely, CaMV aphid transmission Helper Component protein and virion associated protein. Aphid stylets are made of cuticle and little is known about the structure of cuticle protein of this insect group. The present study reports the molecular modeling of the structures of Myzus persicae aphid stylet's cuticular protein (MpsCP) and cauliflower mosaic virus aphid transmission Helper component protein (CaMV HCP). Protein-protein docking studies and molecular dynamics simulations are performed to establish the mode of binding of MpsCP with CaMV HCP. Molecular docking and molecular dynamics investigations of terpenoids Annosquamosin-A from Annona squamosa complex with CaMV transmitting aphid M. persicae stylet's cuticular protein revealed their means of interaction perhaps relates to restrain viral binding and transmission. QM/MM optimization of mesoporous silica nanopores composite with Annosquamosin-A for smart and safe delivery of bioactive is carried out to study their electronic parameters such as heat of formation, total energy, electronic energy, Ionization potential, Highest Occupied Molecular Orbital, Lowest Un-occupied Molecular Orbital and energy gaps.

Formation of large viroplasms and virulence of Cauliflower mosaic virus in turnip plants depend on the N-terminal EKI sequence of viral protein TAV.

Cauliflower mosaic virus (CaMV) TAV protein (TransActivator/Viroplasmin) plays a pivotal role during the infection cycle since it activates translation reinitiation of viral polycistronic RNAs and suppresses RNA silencing. It is also the major component of cytoplasmic electron-dense inclusion bodies (EDIBs) called viroplasms that are particularly evident in cells infected by the virulent CaMV Cabb B-JI isolate. These EDIBs are considered as virion factories, vehicles for CaMV intracellular movement and reservoirs for CaMV transmission by aphids. In this study, focused on different TAV mutants in vivo, we demonstrate that three physically separated domains collectively participate to the formation of large EDIBs: the N-terminal EKI motif, a sequence of the MAV domain involved in translation reinitiation and a C-terminal region encompassing the zinc finger. Surprisingly, EKI mutant TAVm3, corresponding to a substitution of the EKI motif at amino acids 11–13 by three alanines (AAA), which completely abolished the formation of large viroplasms, was not lethal for CaMV but highly reduced its virulence without affecting the rate of systemic infection. Expression of TAVm3 in a viral context led to formation of small irregularly shaped inclusion bodies, mild symptoms and low levels of viral DNA and particles accumulation, despite the production of significant amounts of mature capsid proteins. Unexpectedly, for CaMV-TAVm3 the formation of viral P2-containing electron-light inclusion body (ELIB), which is essential for CaMV aphid transmission, was also altered, thus suggesting an indirect role of the EKI tripeptide in CaMV plant-to-plant propagation. This important functional contribution of the EKI motif in CaMV biology can explain the strict conservation of this motif in the TAV sequences of all CaMV isolates.

Stabilization of cauliflower mosaic virus P3 tetramer by covalent linkage.

Cauliflower mosaic virus (CaMV) open reading frame (ORF) III encodes a 15 kDa protein (P3) that is indispensable for viral infectivity. Although P3 has been shown to be a prerequisite for CaMV aphid transmission, its role in viral replication remains unknown. We previously showed that P3 forms a tetramer in planta and that P3 tetramer co-sediments with viral coat protein on sucrose gradient centrifugation, suggesting that a tetramer may be the functional form of P3. We presumed that disulfide bonds were involved in tetramer formation because 1) the tetramer was detected by Western blotting after electrophoresis under non-reducing conditions, and 2) the cysteine-X-cysteine motif is well conserved in CaMV P3 and P3 homologues among Caulimoviruses. Therefore we mutated either or both of the cysteine residues of CaMV P3. The mutant viruses were infectious and accumulated to a similar extent as the wild-type. An analysis of mutant proteins confirmed that the wild-type P3 molecules in the tetramer are covalently bound with one another through disulfide bonds. It was also suggested that mutant proteins are less stable than wild-type protein in planta. Furthermore, sedimentation study suggested that the disulfide bonds are involved in stable association of P3 with CaMV virions or virion-like particles, or both. The mutant viruses could be transmitted by aphids. These results suggested that the covalent bonds in P3 tetramer are dispensable for biological activity of P3 under experimental situations and may have some biological significance in natural infection in the field.

Interaction between the aphid transmission factor and virus particles is a part of the molecular mechanism of cauliflower mosaic virus aphid transmission.

Cauliflower mosaic virus (CaMV) aphid transmission factor (ATF or P18) is presumed to interact with both virus particles and vector mouthparts, thereby mediating virus aphid transmission. We developed a protein-protein binding assay and our results clearly show that virus particles bind strongly and specifically to P18 whether P18 was obtained from plants, a baculovirus expression system, or the pGEX-3X Escherichia coli expression system. We overproduced, using the pGEX-3X expression system, various fragments of P18 and thereby demonstrated that the C-terminal 31 amino acid residues are responsible for the interaction. Using PCR-based mutagenesis, 2 amino acid residues essential for interaction were identified. Point substitutions (amino acids 157 from Ile to Asn or 159 from Gly to Ser) were sufficient to abolish the interaction, whereas another mutation (amino acid 158 from Ile to Ser) had no effect on P18 virus binding. We evaluated whether there was a correlation between the ability of P18 to interact with CaMV particles and its biological activity. Aphid transmission assays were carried out and we demonstrated that the loss of the virus binding capacity had a dramatic effect on the ability of P18 to mediate aphid transmission. Thus, our results suggest that binding between P18 and virus particles is likely to be one of the molecular mechanisms involved in CaMV aphid transmission.

Paracrystalline Structure of Cauliflower Mosaic Virus Aphid Transmission Factor Produced Both in Plants and in a Heterologous System and Relationship with a Solubilized Active Form

Cauliflower mosaic virus (CaMV) aphid transmission factor (ATF), produced in a baculovirus expression system, forms paracrystalline structures, as demonstrated by electron microscopic observations. Similar paracrystals were also found in CaMV-infected plants, using immunogold techniques, thus providing the first evidence of such a structure for the CaMV ATF (P18). We demonstrated that the paracrystals can be solubilized to provide an active form of the CaMV ATF which can also be reverted into the paracrystalline aggregated form. This suggests that the paracrystalline structures might act as a source of active CaMV ATF or be the form in which it is stored within the infected cells. A point mutation within the CaMV gene II (which encodes the ATF) leads to the loss of both the paracrystalline structures and the ATF activity. Hence, the paracrystalline structure seems to be a feature of the native (unmodified) CaMV ATF.

Cauliflower mosaic virus aphid transmission factor protein is expressed in cells infected with some aphid nontransmissible isolates

An antibody raised to the gel-purified product (p18) of cauliflower mosaic virus (CaMV) open reading frame (ORF) II was used in conjunction with sensitive serological techniques to detect p18 in inclusion body preparations from turnip tissue infected with the aphid transmissible (AT +) isolate Cabb B-JI. p18 was not detected when tissue was infected with the aphid nontransmissible (AT −) isolate CM4-184 that contains a large deletion in ORF II. However, inclusion body preparations from tissue infected with the AT − isolates Campbell and CM1841 contained p18. Hence, the AT phenotype is not directly correlated with the presence or absence of the ORF II product, as was previously indicated when less sensitive techniques were used for analysis.

Cauliflower Mosaic Virus Aphid Transmission Facilitated by Transmission Factors from Other Caulimoviruses

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Cauliflower Mosaic Virus Aphid Transmission Facilitated by Transmission Factors from Other Caulimoviruses

SUMMARY Feeding aphids on plants infected with carnation etched ring virus (CERV) or figwort mosaic virus (FMV) prior to feeding on plants infected with aphid non-transmissible isolates of cauliflower mosaic virus (CaMV) facilitated transmission. This indicates that CERV and FMV code for aphid transmission factors which can substitute for the defective factors in the non-transmissible CaMV isolates.

Helper component for aphid transmission encoded by region II of cauliflower mosaic virus DNA

A deletion mutant of cauliflower mosaic virus (CaMV) isolate NY8153 deficient in aphid transmissibility was constructed by BAL-31 exonuclease treatment of XhoI linearized pCMS31 (a plasmid containing the entire CaMV genome cloned in the SalI site of pBR322), followed by ligation. The resulting mutant, pSA103, lacked about 100 by from the putative protein-coding region II of CaMV DNA. In turnips there was no difference in the number and appearance of starch lesions or hybridization lesions, or in the nature of symptoms in systemically infected leaves induced by pSA103 DNA, pCMS31 DNA, and NY8153 DNA. Systemic symptoms appeared later in plants infected with pSA103 (27 ± 2 days) than in those infected with the parental pCMS31 DNA (19 ± 2 days). Aphids fed on virus SA103-infected mustard plants were unable to transmit the virus to healthy plants while 30–70% transmission was observed from plants infected by the parent virus. Virions extracted from turnips infected with the mutant had DNA still containing the deletion. In addition, the single-stranded discontinuity in region II of NY8153 DNA was missing in this DNA. The results suggest that region II codes for a “helper component” required for aphid transmission of CaMV.

Aphid transmission of cauliflower-mosaic virus

does not appear. First page follows. Summary ELEVEN SPECIES OF APHIDS that do not multiply on cauliflower plants in nature were demonstrated to be vectors of the cauliflower-mosaic virus, five of them being more efficient vectors than three aphid species that do breed on cauliflower. Of three aphid species that breed on cauliflower under natural conditions, the cabbage aphid, Brevicoryne brassicae (Linnaeus), was a more efficient vector of the virus (in single-insect tests) than the turnip or false cabbage aphid, Rhopalosiphum pseudobrassicae (Davis), and the green peach aphid, Myzus persicae (Sulzer). Natural infectivity of the cabbage aphid was demonstrated. None of twenty-one varieties of cauliflower experimentally infected with the virus by the cabbage and green peach aphids was resistant to the disease. Mechanical inoculation was more efficient than transmission by the cabbage, turnip, or green peach aphid. In tests with hourly transfers, most transmissions occurred within 2 hours after the aphids had fed on a mosaic-infected plant. Aphids acquired the virus in 15 to 25 minutes on a diseased plant (5 to 10 minutes actual feeding time) and transmitted it in feeding periods as short as 5 or 10 minutes. In tests with short feeding periods, most of the transmissions occurred during the first 10 minutes after transfer from the infected plant, but an occasional aphid transmitted the disease during the second or third 10-minute period. The cabbage and green peach aphids recovered the virus from inoculated cauliflower plants many days before the first symptom appeared. Mechanical inoculation of healthy cauliflower seedlings and annual stock plants with the centrifuged virus extract from crushed, infective cabbage aphids produced 18 per cent infection.