Nicotiana benthamiana protein, NbPCIP1, interacting with Potato virus X coat protein plays a role as susceptible factor for viral infection (original) (raw)
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Experiments were conducted to compare the plasmodesmal transport activities of Potato virus X (PVX) TGBp1 and coat protein (CP) in several plant species. Microinjection experiments indicated that TGBp1 gates plasmodesmata in Nicotiana tabacum leaves. These results support previous microinjection studies indicating that TGBp1 gates plasmodesmata in Nicotiana benthamiana and Nicotiana clevelandii leaves. To study protein movement, plasmids expressing the green fluorescent protein (GFP) gene fused to the PVX TGBp1 or CP genes were biolistically bombarded to leaves taken from four different PVX host species. GFP/TGBp1 moved between adjacent cells in N. tabacum, N. clevelandii, N. benthamiana, and Lycopersicon esculentum, whereas GFP/CP moved only in N. benthamiana leaves. Mutations m12 and m13 were introduced into the TGBp1 gene and both mutations eliminated TGBp1 ATPase active site motifs, inhibited PVX movement, reduced GFP/TGBp1 cell-to-cell movement in N. benthamiana leaves, and eliminated GFP/TGBp1 movement in N. tabacum, N. clevelandii, and L. esculentum leaves. GFP/TGBp1m13 formed aggregates in tobacco cells. The ability of GFP/CP and mutant GFP/TGBp1 fusion proteins to move in N. benthamiana and not in the other PVX host species suggests that N. benthamiana plants have a unique ability to promote protein intercellular movement.
Journal of Virology, 2007
The capsid protein (CP) of potyviruses is required for various steps during plant infection, such as virion assembly, cell-to-cell movement, and long-distance transport. This suggests a series of compatible interactions with putative host factors which, however, are largely unknown. By using the yeast two-hybrid system the CP from Potato virus Y (PVY) was found to interact with a novel subset of DnaJ-like proteins from tobacco, designated NtCPIPs. Mutational analysis identified the CP core region, previously shown to be essential for virion formation and plasmodesmal trafficking, as the interacting domain. The ability of NtCPIP1 and NtCPIP2a to associate with PVY CP could be confirmed in vitro and was additionally verified in planta by bimolecular fluorescence complementation.
Molecular Plant Pathology, 2012
Potato virus X coat protein is necessary for both cell-to-cell and phloem transfer, but it has not been clarified definitively whether it is needed in both movement phases solely as a component of the assembled particles or also of differently structured ribonucleoprotein complexes. To clarify this issue, we studied the infection progression of a mutant carrying an N-terminal deletion of the coat protein, which was used to construct chimeric virus particles displaying peptides selectively affecting phloem transfer or cell-to-cell movement. Nicotiana benthamiana plants inoculated with expression vectors encoding the wild-type, mutant and chimeric viral genomes were examined by microscopy techniques. These experiments showed that coat protein-peptide fusions promoting cell-to-cell transfer only were not competent for virion assembly, whereas long-distance movement was possible only for coat proteins compatible with virus particle formation. Moreover, the ability of the assembled PVX to enter and persist into developing xylem elements was revealed here for the first time.
Plant Biosystems, 2010
The cytopathology of a Potato virus X (PVX) recombinant variant (encoding as fusion of an epitope of immunological interest with the N‐terminus of the coat protein, PVXSmaP18DD) has been compared with that induced by the wild‐type virus (PVX wt) in Nicotiana benthamiana plants. Both PVX wt and PVXSmaP18DD caused similar ultrastructural alterations, characterized by the presence of laminated inclusion components and bulk virus accumulations in mesophyll cells. However, some striking differences were observed not only in the morphology of these accumulations (typically ordered in PVX wt infection and disordered in PVXSmaP18DD infection) but also because the chimeric virus caused peculiar alterations in chloroplasts structure. Abbreviations: CP, coat protein; d.p.i., days post inoculation; LIC, laminated inclusion components; PVX, Potato virus X
Virology, 2002
The requirements for intercellular movement of Potato virus X (PVX) 12K, 8K, and coat proteins (CP) differed in two Nicotiana spp. Plasmids containing the green fluorescent protein (GFP) gene fused to PVX 12K, 8K, or CP genes were bombarded to tobacco leaves. Protein movement was observed in N. benthamiana but not N. tabacum leaves. GFP:12K and GFP:8K moved cell-to-cell in 25K-expressing transgenic N. tabacum source but not sink leaves. In N. tabacum, GFP:12K and GFP:8K intercellular movement depends on the 25K and leaf developmental stage. Leaves were bombarded using two biolistic delivery methods and the results were similar indicating that movement of GFP:12K or GFP:8K is independent of the delivery system. Mutations in 12K, 8K, and CP genes within the PVX genome inhibited viral intercellular movement in both Nicotiana spp. Thus plasmodesmata gating is not an essential function of these proteins for virus cell-to-cell movement. These proteins likely provide additional activities ...
Virology, 2003
Potato virus X (PVX) TGBp1, TGBp2, TGBp3, and coat protein are required for virus cell-to-cell movement. Plasmids expressing GFP fused to TGBp2 were bombarded to leaf epidermal cells and GFP:TGBp2 moved cell to cell in Nicotiana benthamiana leaves but not in Nicotiana tabacum leaves. GFP:TGBp2 movement was observed in TGBp1-transgenic N. tabacum, indicating that TGBp2 requires TGBp1 to promote its movement in N. tabacum. In this study, GFP:TGBp2 was detected in a polygonal pattern that resembles the endoplasmic reticulum (ER) network. Amino acid sequence analysis revealed TGBp2 has two putative transmembrane domains. Two mutations separately introduced into the coding sequences encompassing the putative transmembrane domains within the GFP:TGBp2 plasmids and PVX genome, disrupted membrane binding of GFP:TGBp2, inhibited GFP:TGBp2 movement in N. benthamiana and TGBp1-expressing N. tabacum, and inhibited PVX movement. A third mutation, lying outside the transmembrane domains, had no effect on GFP:TGBp2 ER association or movement in N. benthamiana but inhibited GFP:TGBp2 movement in TGBp1-expressing N. tabacum and PVX movement in either Nicotiana species. Thus, ER association of TGBp2 may be required but not be sufficient for virus movement. TGBp2 likely provides an activity for PVX movement beyond ER association.
Molecular Plant-microbe Interactions, 2006
The cylindrical inclusion (CI) protein of potyviruses is involved in virus replication and cell-to-cell movement. These two processes should rely on multiple plant-virus interactions; however, little is known about the host factors that are involved in, or that may interfere with, CI functions. By using a yeast two-hybrid system, the CI protein from Plum pox virus (PPV) was found to interact with the photosystem I PSI-K protein, the product of the gene psaK, of Nicotiana benthamiana.
Virus Research, 2007
The N-terminal (NT) regions of particular protein-coding sequences are generally used for in-frame insertion of heterologous open reading frames (ORFs) in potyviral vectors for protein expression in plants. An infectious cDNA clone of Turnip mosaic virus (TuMV) isolate YC5 was engineered at the generally used NT regions of HC-Pro and CP, and other possibly permissive sites to investigate their effectiveness to express the GFP (jellyfish green fluorescent protein) and Der p 5 (allergen from the dust mite, Dermatophagoides pteronyssinus) ORFs. The results demonstrated the permissiveness of the NT regions of P3, CIP and NIb to carry the ORFs and express the translates as part of the viral polyprotein, the processing of which released free-form proteins in the host cell milieu. However, these sites varied in their permissiveness to retain the ORFs intact and hence affect the heterologous protein expression. Moreover, strong influence of the inserted ORF and host plants in determining the permissiveness of a viral genomic context to stably carry the alien ORFs and hence to support their prolonged expression was also noticed. In general, the engineered sites were relatively more permissive to the GFP ORF than to the Der p 5 ORF. Among the hosts, the local lesion host, Chenopodium quinoa Willd. showed the highest extent of support to TuMV to stably carry the heterologous ORFs at the engineered sites and the protein expression therefrom. Among the systemic hosts, Nicotiana benthamiana Domin proved more supportive to TuMV to carry and express the heterologous ORFs than the Brassica hosts, whereas the protein expression levels were significantly higher and more stable in the plants of Brassica campestris L. var. chinensis and B. campestris L. var. ching-geeng than those in the plants of B. juncea L. and B. campestris L. var. pekinensis.