scholarly journals Independent Expression of the First Two Triple Gene Block Proteins of Beet Necrotic Yellow Vein Virus Complements Virus Defective in the Corresponding Gene but Expression of the Third Protein Inhibits Viral Cell-to-Cell Movement

1997 ◽  
Vol 10 (2) ◽  
pp. 240-246 ◽  
Author(s):  
Claudine Bleykasten-Grosshans ◽  
H. Guilley ◽  
S. Bouzoubaa ◽  
K. E. Richards ◽  
G. Jonard
Keyword(s):  
Triple Gene Block ◽  
Cell Movement ◽  
Viral Rna ◽  
Yellow Vein ◽  
Mixed Infections ◽  
Subgenomic Rna ◽  
Gene Block ◽  
The Third ◽  
Local Lesion Formation ◽  
P15 Gene

Cell-to-cell movement of beet necrotic yellow vein furovirus is controlled by three slightly overlapping genes on RNA 2 called the triple gene block (TGB) encoding, in order, P42, P13, and P15. Synthesis of P42 is directed by subgenomic RNA 2suba while synthesis of both P13 and P15 is probably directed by a dicistronic subgenomic RNA, 2subb. For complementation experiments, each TGB protein gene was inserted into a “replicon” derived from viral RNA 3. In mixed infections, the replicons expressing P42 and P13 complemented RNA 2 mutants defective in the corresponding gene. A P15-containing replicon did not complement a P15-defective RNA 2 but complementation was observed with a dicistronic replicon containing the P15 gene placed behind the P13 gene. In mixed infections with wild-type viral RNAs, the P15-containing replicon did not inhibit viral RNA replication in protoplasts but blocked local lesion formation on leaves. Infection of leaves was also inhibited by an RNA3-derived replicon containing the third TGB gene from another furovirus, peanut clump virus. The results are consistent with a model in which viral cell-to-cell movement requires production of appropriate relative amounts of P13 and P15, and their expression from a dicistronic subgenomic RNA provides a mechanism for coordinating their synthesis.

scholarly journals P42 Movement Protein of Beet necrotic yellow vein virus Is Targeted by the Movement Proteins P13 and P15 to Punctate Bodies Associated with Plasmodesmata

2000 ◽  
Vol 13 (5) ◽  
pp. 520-528 ◽  
Author(s):  
M. Erhardt ◽  
M. Morant ◽  
C. Ritzenthaler ◽  
C. Stussi-Garaud ◽  
H. Guilley ◽  
...  
Keyword(s):  
Cell Wall ◽  
Fluorescent Protein ◽  
Triple Gene Block ◽  
Point Mutations ◽  
Cell Movement ◽  
Leading Edge ◽  
Yellow Vein ◽  
Body Formation ◽  
Movement Proteins ◽  
Gene Block

Cell-to-cell movement of Beet necrotic yellow vein virus (BNYVV) is driven by a set of three movement proteins—P42, P13, and P15—organized into a triple gene block (TGB) on viral RNA 2. The first TGB protein, P42, has been fused to the green fluorescent protein (GFP) and fusion proteins between P42 and GFP were expressed from a BNYVV RNA 3-based replicon during virus infection. GFP-P42, in which the GFP was fused to the P42 N terminus, could drive viral cell-to-cell movement when the copy of the P42 gene on RNA 2 was disabled but the C-terminal fusion P42-GFP could not. Confocal microscopy of epidermal cells of Chenopodium quinoa near the leading edge of the infection revealed that GFP-P42 localized to punctate bodies apposed to the cell wall whereas free GFP, expressed from the replicon, was distributed uniformly throughout the cytoplasm. The punctate bodies sometimes appeared to traverse the cell wall or to form pairs of disconnected bodies on each side. The punctate bodies co-localized with callose, indicating that they are associated with plasmodesmata-rich regions such as pit fields. Point mutations in P42 that inhibited its ability to drive cell-to-cell movement also inhibited GFP-P42 punctate body formation. GFP-P42 punctate body formation was dependent on expression of P13 and P15 during the infection, indicating that these proteins act together or sequentially to localize P42 to the plasmodesmata.

scholarly journals Cell-to-Cell Movement of Beet Necrotic Yellow Vein Virus: I. Heterologous Complementation Experiments Provide Evidence for Specific Interactions Among the Triple Gene Block Proteins

1998 ◽  
Vol 11 (7) ◽  
pp. 618-625 ◽  
Author(s):  
Emmanuelle Lauber ◽  
Claudine Bleykasten-Grosshans ◽  
M. Erhardt ◽  
S. Bouzoubaa ◽  
G. Jonard ◽  
...  
Keyword(s):  
Mosaic Virus ◽  
Triple Gene Block ◽  
Chenopodium Quinoa ◽  
Alfalfa Mosaic Virus ◽  
Cell Movement ◽  
Yellow Vein ◽  
Specific Interactions ◽  
In Planta ◽  
Movement Proteins ◽  
Gene Block

Cell-to-cell movement of beet necrotic yellow vein virus (BNYVV) requires three proteins encoded by a triple gene block (TGB) on viral RNA 2. A BNYVV RNA 3-derived replicon was used to express movement proteins of other viruses and the ability of these proteins to functionally substitute for the BNYVV TGB proteins was tested by coinoculation of TGB-defective BNYVV with the various replicons to Chenopodium quinoa. Trans-heterocomplementation was successful with the movement protein (P30) of tobacco mosaic virus but not with the tubule-forming movement proteins of alfalfa mosaic virus and grapevine fanleaf virus. Trans-complementation of BNYVV movement was also observed when all three TGB proteins of the distantly related peanut clump virus were supplied together but not when they were substituted for their BNYVV counterparts one by one. When P30 was used to drive BNYVV movement in trans, accumulation of the first TGB protein of BNYVV was adversely affected by null mutations in the second and third TGB proteins. Taken together, these results suggest that highly specific interactions among cognate TGB proteins are important for their function and/or stability in planta.

Small RNA and transcriptome sequencing of a symptomatic peony plant reveals mixed infections with novel viruses

Plant Disease ◽  
2021 ◽  
Author(s):  
Anning Jia ◽  
Chenge Yan ◽  
Hang Yin ◽  
Rui Sun ◽  
Fei Xia ◽  
...  
Keyword(s):  
Phylogenetic Analysis ◽  
Small Rna ◽  
High Throughput Sequencing ◽  
Viral Infections ◽  
Triple Gene Block ◽  
Field Investigation ◽  
Mixed Infections ◽  
Species Classification ◽  
Gene Block ◽  
Sequence Identity

To identify the viruses in tree peony plants associated with the symptoms of yellowing, leaf rolling, stunted growth, and decline, high-throughput sequencing of small RNA and mRNA was conducted from a single symptomatic plant. Bioinformatic analyses and reconstruction of viral genomes indicated mixed viral infections involving cycas necrotic stunt virus (CNSV), apple stem grooving virus (ASGV), lychnis mottle virus (LycMoV), grapevine line pattern virus (GLPV), and three new viruses designated as peony yellowing-associated citrivirus (PYaCV, Citrivirus in Betaflexiviridae), peony betaflexivirus 1 (PeV1, unclassified in Betaflexiviridae), and peony leafroll-associated virus (PLRaV, Ampelovirus in Closteroviridae). PYaCV was 8,666 nucleaotides (nt) in length, comprising three open reading frames (ORFs) and shared 63.8–75.9% nucleotide sequence identity with citrus leaf blotch virus (CLBV) isolates. However, the ORF encoding the replication-associated protein (REP) shared 57% and 52% sequence identities at the nt and amino acid (aa) level, respectively, with other reported CLBV isolates, which were below the criterion for species classification within the family Betaflexiviridae. Recombination analysis identified putative recombination sites in PYaCV, which originated from CLBV. PeV1, only identified from the transcriptome data, was 8,124 nt in length with five ORFs encoding the REP (ORF1), triple gene block (TGB, ORF2–4) and coat protein (CP, ORF5) proteins. Phylogenetic analysis and sequence comparison showed that PeV1 clustered with an unassigned member, the garlic yellow mosaic-associated virus (GYMaV) within the Betaflexiviridae family, into a separate clade. Partial genome sequence analysis of PLRaV (12,545 nt) showed it contained seven ORFs encoding the partial polyprotein 1a, the RNA-dependent RNA polymerase (RdRp), two small hydrophobic proteins p11 and p6, HSP70h, p55, and a CP duplicate, which shared low aa sequence identity with Closteroviridae family members. Phylogenetic analysis based on the aa sequences of RdRp or HSP70h indicated that PLRaV clustered with grapevine leafroll-associated virus 1 (GLRaV-1) and GLRaV-13 in the Ampelovirus genus. Field investigation confirmed the wide distribution of these viruses, causing mixed infections of peony plants in Beijing.

scholarly journals Triple Gene Block Protein Interactions Involved in Movement of Barley Stripe Mosaic Virus

2008 ◽  
Vol 82 (10) ◽  
pp. 4991-5006 ◽  
Author(s):  
Hyoun-Sub Lim ◽  
Jennifer N. Bragg ◽  
Uma Ganesan ◽  
Diane M. Lawrence ◽  
Jialin Yu ◽  
...  
Keyword(s):  
Mosaic Virus ◽  
Triple Gene Block ◽  
Cell Movement ◽  
Wild Type Virus ◽  
Barley Stripe Mosaic Virus ◽  
Wild Type ◽  
In Planta ◽  
Gene Block ◽  
Physical Interactions

ABSTRACT Barley stripe mosaic virus (BSMV) encodes three movement proteins in an overlapping triple gene block (TGB), but little is known about the physical interactions of these proteins. We have characterized a ribonucleoprotein (RNP) complex consisting of the TGB1 protein and plus-sense BSMV RNAs from infected barley plants and have identified TGB1 complexes in planta and in vitro. Homologous TGB1 binding was disrupted by site-specific mutations in each of the first two N-terminal helicase motifs but not by mutations in two C-terminal helicase motifs. The TGB2 and TGB3 proteins were not detected in the RNP, but affinity chromatography and yeast two-hybrid experiments demonstrated that TGB1 binds to TGB3 and that TGB2 and TGB3 form heterologous interactions. These interactions required the TGB2 glycine 40 and the TGB3 isoleucine 108 residues, and BSMV mutants containing these amino acid substitution were unable to move from cell to cell. Infectivity experiments indicated that TGB1 separated on a different genomic RNA from TGB2 and TGB3 could function in limited cell-to-cell movement but that the rates of movement depended on the levels of expression of the proteins and the contexts in which they are expressed. Moreover, elevated expression of the wild-type TGB3 protein interfered with cell-to-cell movement but movement was not affected by the similar expression of a TGB3 mutant that fails to interact with TGB2. These experiments suggest that BSMV movement requires physical interactions of TGB2 and TGB3 and that substantial deviation from the TGB protein ratios expressed by the wild-type virus compromises movement.

scholarly journals Cell-to-Cell Movement of Potato virus X: The Role of p12 and p8 Encoded by the Second and Third Open Reading Frames of the Triple Gene Block

2001 ◽  
Vol 14 (10) ◽  
pp. 1158-1167 ◽  
Author(s):  
Atsushi Tamai ◽  
Tetsuo Meshi
Keyword(s):  
Potato Virus ◽  
Fluorescent Protein ◽  
Microprojectile Bombardment ◽  
Triple Gene Block ◽  
Cell Movement ◽  
Potato Virus X ◽  
Open Reading Frames ◽  
Gene Block ◽  
Infected Cells ◽  
Green Fluorescent

Potato virus X (PVX) requires three proteins, p25, p12, and p8, encoded by the triple gene block plus the coat protein (CP) for cell-to-cell movement. When each of these proteins was co-expressed with a cytosolic green fluorescent protein (GFP) in the epidermal cells of Nicotiana benthamiana by the microprojectile bombardment-mediated gene delivery method, only p12 enhanced diffusion of co-expressed GFP, indicating an ability to alter plasmodesmal permeability. p25, p12, and CP, expressed transiently in the initially infected cells, transcomplemented the corresponding movement-defective mutants to spread through two or more cell boundaries. Thus, these proteins probably move from cell to cell with the genomic RNA. In contrast, p8 only functioned intracellularly and was not absolutely required for cell-to-cell movement. Since overexpression of p12 overcame the p8 deficiency, p8 appears to facilitate the functioning of p12, presumably by mediating its intracellular trafficking. Considering the likelihood that p12 and p8 are membrane proteins, it is suggested that intercellular as well as intracellular movement of PVX involves a membrane-mediated process.

scholarly journals Interactions of the TGB1 Protein during Cell-to-Cell Movement of Barley Stripe Mosaic Virus

2001 ◽  
Vol 75 (18) ◽  
pp. 8712-8723 ◽  
Author(s):  
Diane M. Lawrence ◽  
A. O. Jackson
Keyword(s):  
Subcellular Localization ◽  
Mosaic Virus ◽  
Fluorescent Protein ◽  
Triple Gene Block ◽  
Cell Movement ◽  
Barley Stripe Mosaic Virus ◽  
Helicase Domain ◽  
Chenopodium Amaranticolor ◽  
Gene Block ◽  
Green Fluorescent

ABSTRACT We have recently used a green fluorescent protein (GFP) fusion to the γb protein of Barley stripe mosaic virus (BSMV) to monitor cell-to-cell and systemic virus movement. The γb protein is involved in expression of the triple gene block (TGB) proteins encoded by RNAβ but is not essential for cell-to-cell movement. The GFP fusion appears not to compromise replication or movement substantially, and mutagenesis experiments demonstrated that the three most abundant TGB-encoded proteins, βb (TGB1), βc (TGB3), and βd (TGB2), are each required for cell-to-cell movement (D. M. Lawrence and A. O. Jackson, Mol. Plant Pathol. 2:65–75, 2001). We have now extended these analyses by engineering a fusion of GFP to TGB1 to examine the expression and interactions of this protein during infection. BSMV derivatives containing the TGB1 fusion were able to move from cell to cell and establish local lesions in Chenopodium amaranticolor and systemic infections of Nicotiana benthamiana and barley. In these hosts, the GFP-TGB1 fusion protein exhibited a temporal pattern of expression along the advancing edge of the infection front. Microscopic examination of the subcellular localization of the GFP-TGB1 protein indicated an association with the endoplasmic reticulum and with plasmodesmata. The subcellular localization of the TGB1 protein was altered in infections in which site-specific mutations were introduced into the six conserved regions of the helicase domain and in mutants unable to express the TGB2 and/or TGB3 proteins. These results are compatible with a model suggesting that movement requires associations of the TGB1 protein with cytoplasmic membranes that are facilitated by the TGB2 and TGB3 proteins.

scholarly journals Subcellular localization of the Triple Gene Block movement proteins of Beet necrotic yellow vein virus by electron microscopy

Virology ◽  
2005 ◽  
Vol 340 (1) ◽  
pp. 155-166 ◽  
Author(s):  
M. Erhardt ◽  
G. Vetter ◽  
D. Gilmer ◽  
S. Bouzoubaa ◽  
K. Richards ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Subcellular Localization ◽  
Triple Gene Block ◽  
Yellow Vein ◽  
Movement Proteins ◽  
Gene Block ◽  
Block Movement

scholarly journals Cell-to-Cell Movement of Potexviruses: Evidence for a Ribonucleoprotein Complex Involving the Coat Protein and First Triple Gene Block Protein

2000 ◽  
Vol 13 (9) ◽  
pp. 962-974 ◽  
Author(s):  
Tony J. Lough ◽  
Natalie E. Netzler ◽  
Sarah J. Emerson ◽  
Paul Sutherland ◽  
Fiona Carr ◽  
...  
Keyword(s):  
Coat Protein ◽  
Intracellular Transport ◽  
Microprojectile Bombardment ◽  
Triple Gene Block ◽  
Cell Movement ◽  
Limiting Factor ◽  
Ribonucleoprotein Complex ◽  
Gene Block ◽  
Cellular Machinery ◽  
Mode Of Transport

The triple gene block proteins (TGBp1-3) and coat protein (CP) of potexviruses are required for cell-to-cell movement. Separate models have been proposed for inter-cellular movement of two of these viruses, transport of intact virions, or a ribonucleoprotein complex (RNP) comprising genomic RNA, TGBp1, and the CP. At issue therefore, is the form(s) in which RNA transport occurs and the roles of TGBp1-3 and the CP in movement. Evidence is presented that, based on microprojectile bombardment studies, TGBp1 and the CP, but not TGBp2 or TGBp3, are co-translocated between cells with viral RNA. In addition, cell-to-cell movement and encapsidation functions of the CP were shown to be separable, and the rate-limiting factor of potexvirus movement was shown not to be virion accumulation, but rather, the presence of TGBp1-3 and the CP in the infected cell. These findings are consistent with a common mode of transport for potex-viruses, involving a non-virion RNP, and show that TGBp1 is the movement protein, whereas TGBp2 and TGBp3 are either involved in intracellular transport or interact with the cellular machinery/docking sites at the plasmodesmata.

scholarly journals Movement of potexviruses requires species-specific interactions among the cognate triple gene block proteins, as revealed by a trans-complementation assay based on the bamboo mosaic virus satellite RNA-mediated expression system

2006 ◽  
Vol 87 (5) ◽  
pp. 1357-1367 ◽  
Author(s):  
Ming-Kuem Lin ◽  
Chung-Chi Hu ◽  
Na-Sheng Lin ◽  
Ban-Yang Chang ◽  
Yau-Heiu Hsu
Keyword(s):  
Mosaic Virus ◽  
Expression System ◽  
Triple Gene Block ◽  
Ectopic Expression ◽  
Cell Movement ◽  
Specific Interactions ◽  
Satellite Rna ◽  
Complementation Assay ◽  
Gene Block ◽  
Species Specific

The intra- and intercellular transport of potexviruses require interactions among viral RNA, coat protein and elements of the triple gene block proteins (TGBps). In this study, the requirement of bamboo mosaic virus (BaMV) TGBps for movement functions and the compatibilities with those of two potexviruses, Potato virus X (PVX) and Foxtail mosaic virus (FoMV), were examined using a satellite RNA-mediated trans-complementation assay system. Single or multiple TGBps of BaMV, PVX and FoMV were expressed from BaMV satellite RNA (satBaMV RNA) vectors to complement the functions of green fluorescent protein-tagged, movement-defective BaMV with mutation(s) in the matching gene(s). It was found that individual BaMV TGBps expressed from the satellite vector could function normally in trans, whereas bi-gene BaMV TGBp constructs in which the expression of TGBp3 might be impaired and individual TGBp genes from PVX or FoMV could not complement the movement functions of the defective helper viruses. Furthermore, alterations of the ratio among TGBps by ectopic expression of individual components of TGBps from satBaMV RNA vectors did not affect the cell-to-cell movement capabilities of wild-type BaMV significantly. The results indicate that species-specific interactions among movement proteins are obligatory for the cell-to-cell movement of BaMV and possibly other potexviruses.

scholarly journals Mutation of a chloroplast-targeting signal in Alternanthera mosaic virus TGB3 impairs cell-to-cell movement and eliminates long-distance virus movement

2010 ◽  
Vol 91 (8) ◽  
pp. 2102-2115 ◽  
Author(s):  
Hyoun-Sub Lim ◽  
Anna Maria Vaira ◽  
Hanhong Bae ◽  
Jennifer N. Bragg ◽  
Steven E. Ruzin ◽  
...  
Keyword(s):  
Mosaic Virus ◽  
Critical Role ◽  
Triple Gene Block ◽  
Cell Movement ◽  
Potato Virus X ◽  
Start Codon ◽  
Long Distance ◽  
Mesophyll Cells ◽  
Chloroplast Targeting ◽  
Gene Block

Cell-to-cell movement of potexviruses requires coordinated action of the coat protein and triple gene block (TGB) proteins. The structural properties of Alternanthera mosaic virus (AltMV) TGB3 were examined by methods differentiating between signal peptides and transmembrane domains, and its subcellular localization was studied by Agrobacterium-mediated transient expression and confocal microscopy. Unlike potato virus X (PVX) TGB3, AltMV TGB3 was not associated with the endoplasmic reticulum, and accumulated preferentially in mesophyll cells. Deletion and site-specific mutagenesis revealed an internal signal VL(17,18) of TGB3 essential for chloroplast localization, and either deletion of the TGB3 start codon or alteration of the chloroplast-localization signal limited cell-to-cell movement to the epidermis, yielding a virus that was unable to move into the mesophyll layer. Overexpression of AltMV TGB3 from either AltMV or PVX infectious clones resulted in veinal necrosis and vesiculation at the chloroplast membrane, a cytopathology not observed in wild-type infections. The distinctive mesophyll and chloroplast localization of AltMV TGB3 highlights the critical role played by mesophyll targeting in virus long-distance movement within plants.

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