scholarly journals Subcellular Localization of the Barley Stripe Mosaic Virus Triple Gene Block Proteins

2009 ◽  
Vol 83 (21) ◽  
pp. 11413-11413 ◽  
Author(s):  
Hyoun-Sub Lim ◽  
Jennifer N. Bragg ◽  
Uma Ganesan ◽  
Steven Ruzin ◽  
Denise Schichnes ◽  
...  
Keyword(s):  
Subcellular Localization ◽  
Mosaic Virus ◽  
Triple Gene Block ◽  
Barley Stripe Mosaic Virus ◽  
Gene Block

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 Barley Stripe Mosaic Virus Triple Gene Block Proteins

2009 ◽  
Vol 83 (18) ◽  
pp. 9432-9448 ◽  
Author(s):  
Hyoun-Sub Lim ◽  
Jennifer N. Bragg ◽  
Uma Ganesan ◽  
Steven Ruzin ◽  
Denise Schichnes ◽  
...  
Keyword(s):  
Subcellular Localization ◽  
Mosaic Virus ◽  
Protein Interactions ◽  
Close Association ◽  
Triple Gene Block ◽  
Single Amino Acid ◽  
Barley Stripe Mosaic Virus ◽  
Subcellular Targeting ◽  
Site Specific ◽  
Gene Block

ABSTRACT Barley stripe mosaic virus (BSMV) spreads from cell to cell through the coordinated actions of three triple gene block (TGB) proteins (TGB1, TGB2, and TGB3) arranged in overlapping open reading frames (ORFs). Our previous studies (D. M. Lawrence and A. O. Jackson, J. Virol. 75:8712-8723, 2001; D. M. Lawrence and A. O. Jackson, Mol. Plant Pathol. 2:65-75, 2001) have shown that each of these proteins is required for cell-to-cell movement in monocot and dicot hosts. We recently found (H.-S. Lim, J. N. Bragg, U. Ganesan, D. M. Lawrence, J. Yu, M. Isogai, J. Hammond, and A. O. Jackson, J. Virol. 82:4991-5006, 2008) that TGB1 engages in homologous interactions leading to the formation of a ribonucleoprotein complex containing viral genomic and messenger RNAs, and we have also demonstrated that TGB3 functions in heterologous interactions with TGB1 and TGB2. We have now used Agrobacterium tumefaciens-mediated protein expression in Nicotiana benthamiana leaf cells and site-specific mutagenesis to determine how TGB protein interactions influence their subcellular localization and virus spread. Confocal microscopy revealed that the TGB3 protein localizes at the cell wall (CW) in close association with plasmodesmata and that the deletion or mutagenesis of a single amino acid at the immediate C terminus can affect CW targeting. TGB3 also directed the localization of TGB2 from the endoplasmic reticulum to the CW, and this targeting was shown to be dependent on interactions between the TGB2 and TGB3 proteins. The optimal localization of the TGB1 protein at the CW also required TGB2 and TGB3 interactions, but in this context, site-specific TGB1 helicase motif mutants varied in their localization patterns. The results suggest that the ability of TGB1 to engage in homologous binding interactions is not essential for targeting to the CW. However, the relative expression levels of TGB2 and TGB3 influenced the cytosolic and CW distributions of TGB1 and TGB2. Moreover, in both cases, localization at the CW was optimal at the 10:1 TGB2-to-TGB3 ratios occurring in virus infections, and mutations reducing CW localization had corresponding effects on BSMV movement phenotypes. These data support a model whereby TGB protein interactions function in the subcellular targeting of movement protein complexes and the ability of BSMV to move from cell to cell.

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 Actin Cytoskeleton and Golgi Involvement in Barley stripe mosaic virus Movement and Cell Wall Localization of Triple Gene Block Proteins

2013 ◽  
Vol 29 (1) ◽  
pp. 17-30 ◽  
Author(s):  
Hyoun-Sub Lim ◽  
Mi Yeon Lee ◽  
Jae Sun Moon ◽  
Jung-Kyung Moon ◽  
Yong-Man Yu ◽  
...  
Keyword(s):  
Cell Wall ◽  
Actin Cytoskeleton ◽  
Mosaic Virus ◽  
Triple Gene Block ◽  
Barley Stripe Mosaic Virus ◽  
Virus Movement ◽  
Gene Block

scholarly journals Barley stripe mosaic virus-encoded proteins triple-gene block 2 and γb localize to chloroplasts in virus-infected monocot and dicot plants, revealing hitherto-unknown roles in virus replication

2006 ◽  
Vol 87 (8) ◽  
pp. 2403-2411 ◽  
Author(s):  
L. Torrance ◽  
G. H. Cowan ◽  
T. Gillespie ◽  
A. Ziegler ◽  
C. Lacomme
Keyword(s):  
Mosaic Virus ◽  
Virus Replication ◽  
Triple Gene Block ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Barley Stripe Mosaic Virus ◽  
Cytoplasmic Inclusions ◽  
Thin Sections ◽  
Gene Block ◽  
Encoded Proteins

Replication of Barley stripe mosaic virus (BSMV), genus Hordeivirus, is thought to be associated with vesicles in proplastids and chloroplasts, but the molecular details of the process and identity of virus proteins involved in establishing the virus replication complexes are unknown. In addition, BSMV encodes a triple-gene block of movement proteins (TGBs) that putatively share functional roles with their counterparts in other hordei-, pomo- and pecluviruses, but detailed information on the intracellular locations of the individual TGBs is lacking. Here, the subcellular localizations of BSMV-encoded proteins TGB2 and γb fused to green or red fluorescent proteins were examined in epidermal cells of Nicotiana benthamiana and barley (Hordeum vulgare ‘Black Hulless’). The fusion proteins were expressed from a BSMV vector or under the control of the cauliflower mosaic virus 35S promoter. The subcellular localizations were studied by confocal laser-scanning microscopy (CLSM). CLSM studies showed that both proteins were recruited to chloroplasts in the presence of viral RNA and that virus RNA, coat protein and γb protein were detected in plastid preparations from infected leaves. Electron microscope images of thin sections of virus-infected leaves revealed abnormal chloroplasts with cytoplasmic inclusions containing virus-like particles. In addition, cellular localizations of BSMV TGB2 suggest subtle differences in function between the hordei-like TGB2 proteins. The results indicate that TGB2 and γb proteins play a previously unknown functional role at the site of virus replication.

scholarly journals Expression of the Barley Stripe Mosaic Virus RNAβ “Triple Gene Block”

Virology ◽  
1996 ◽  
Vol 216 (2) ◽  
pp. 367-379 ◽  
Author(s):  
HUAN ZHOU ◽  
A.O. JACKSON
Keyword(s):  
Mosaic Virus ◽  
Triple Gene Block ◽  
Barley Stripe Mosaic Virus ◽  
Gene Block

Comparison of Barley Stripe Mosaic Virus Strains

2008 ◽  
Vol 63 (3-4) ◽  
pp. 271-276 ◽  
Author(s):  
Elsayed E. Hafez ◽  
Engy E. Abdel Aleem ◽  
Faiza A. Fattouh
Keyword(s):  
Mosaic Virus ◽  
Phylogenetic Trees ◽  
Triple Gene Block ◽  
Northern Blotting ◽  
Barley Stripe Mosaic Virus ◽  
Evolutionary Divergence ◽  
Gene Block ◽  
Virus Particles ◽  
Distinct Cluster ◽  
High Level

BSMV (barley stripe mosaic virus) particles were obtained in a pure state from infected host plant tissues of Hordeum vulgare. The three genomic parities (α, β and γ) were amplified by PCR using specific primers for each particle; each was cloned. Partial sequence of the α, β and γ segments was determined for the Egyptian isolate of barley stripe mosaic virus (BSMV AE1). Alignment of nucleotide sequences with that of other known strains of the virus, BSMV type strains (CV17, ND18 and China), and the generation of phylogenetic trees was performed. A low level of homology was detected comparing 467 bp of the α and 643 bp of the segments to that of the other strains, and thus BSMV α and β segments were in separate clusters. However, 1154 bp of the γ segments of BSMV AE1 showed a high level of homology especially to strain BSMV ND18, as they both formed a distinct cluster. Northern blotting of pure BSMV AE1 virus and H. vulgare-infected tissue were compared using an α ND18 specific probe. Western blotting using antibodies specific for the coat protein (CP) and the triple gene block 1 (TGB1) protein, which are both encoded by the β ND18 segment, still indicated a high level of similarity between proteins produced by BSMV ND18 and AE1. We suggest that the BSMV AE1 isolate is a distinct strain of BSMV which reflects the genetic evolutionary divergence among BSMV strains and members of the Hordeivirus group

Brachypodium distachyon line Bd3-1 resistance is elicited by the barley stripe mosaic virus triple gene block 1 movement protein

2012 ◽  
Vol 93 (12) ◽  
pp. 2729-2739 ◽  
Author(s):  
Mi Yeon Lee ◽  
Lijie Yan ◽  
Florien A. Gorter ◽  
Brian Y. T. Kim ◽  
Yu Cui ◽  
...  
Keyword(s):  
Mosaic Virus ◽  
Double Mutant ◽  
Movement Protein ◽  
Brachypodium Distachyon ◽  
Triple Gene Block ◽  
Barley Stripe Mosaic Virus ◽  
Double Mutation ◽  
Gene Block ◽  
Necrotic Spots ◽  
Systemic Infections

Barley stripe mosaic virus North Dakota 18 (ND18), Beijing (BJ), Xinjiang (XJ), Type (TY) and CV21 strains are unable to infect the Brachypodium distachyon Bd3-1 inbred line, which harbours a resistance gene designated Bsr1, but the Norwich (NW) strain is virulent on Bd3-1. Analysis of ND18 and NW genomic RNA reassortants and RNAβ mutants demonstrates that two amino acids within the helicase motif of the triple gene block 1 (TGB1) movement protein have major effects on their Bd3-1 phenotypes. Resistance to ND18 correlates with an arginine residue at TGB1 position 390 (R390) and a threonine at position 392 (T392), whereas the virulent NW strain contains lysines (K) at both positions. ND18 TGB1 R390K (NDTGB1R390K) and NDTGB1T392K single substitutions, and an NDTGB1R390K,T392K double mutation resulted in systemic infections of Bd3-1. Reciprocal NDTGB1 substitutions into NWTGB1 (NWTGB1K390R and NWTGB1K392T) failed to affect virulence, implying that K390 and K392 compensate for each other. In contrast, an NWTGB1K390R,K392T double mutant exhibited limited vascular movement in Bd3-1, but developed prominent necrotic streaks that spread from secondary leaf veins. This phenotype, combined with the appearance of necrotic spots in certain ND18 mutants, and necrosis and rapid wilting of Bd3-1 plants after BJ strain (BJTGB1K390,T392) inoculations, show that Bd3-1 Bsr1 resistance is elicited by the TGB1 protein and suggest that it involves a hypersensitive response.

scholarly journals A Thioredoxin Domain-Containing Protein Interacts with Pepino mosaic virus Triple Gene Block Protein 1

2018 ◽  
Vol 19 (12) ◽  
pp. 3747
Author(s):  
Matthaios Mathioudakis ◽  
Souheyla Khechmar ◽  
Carolyn Owen ◽  
Vicente Medina ◽  
Karima Ben Mansour ◽  
...  
Keyword(s):  
Mosaic Virus ◽  
Triple Gene Block ◽  
Polyclonal Antiserum ◽  
Post Inoculation ◽  
Mrna Levels ◽  
In Planta ◽  
Pepino Mosaic Virus ◽  
Gene Block ◽  
Triple Gene Block Protein

Pepino mosaic virus (PepMV) is a mechanically-transmitted tomato pathogen of importance worldwide. Interactions between the PepMV coat protein and triple gene block protein (TGBp1) with the host heat shock cognate protein 70 and catalase 1 (CAT1), respectively, have been previously reported by our lab. In this study, a novel tomato interactor (SlTXND9) was shown to bind the PepMV TGBp1 in yeast-two-hybrid screening, in vitro pull-down and bimolecular fluorescent complementation (BiFC) assays. SlTXND9 possesses part of the conserved thioredoxin (TRX) active site sequence (W__PC vs. WCXPC), and TXND9 orthologues cluster within the TRX phylogenetic superfamily closest to phosducin-like protein-3. In PepMV-infected and healthy Nicotiana benthamiana plants, NbTXND9 mRNA levels were comparable, and expression levels remained stable in both local and systemic leaves for 10 days post inoculation (dpi), as was also the case for catalase 1 (CAT1). To localize the TXND9 in plant cells, a polyclonal antiserum was produced. Purified α-SlTXND9 immunoglobulin (IgG) consistently detected a set of three protein bands in the range of 27–35 kDa, in the 1000 and 30,000 g pellets, and the soluble fraction of extracts of healthy and PepMV-infected N. benthamiana leaves, but not in the cell wall. These bands likely consist of the homologous protein NbTXND9 and its post-translationally modified derivatives. On electron microscopy, immuno-gold labelling of ultrathin sections of PepMV-infected N. benthamiana leaves using α-SlTXND9 IgG revealed particle accumulation close to plasmodesmata, suggesting a role in virus movement. Taken together, this study highlights a novel tomato-PepMV protein interaction and provides data on its localization in planta. Currently, studies focusing on the biological function of this interaction during PepMV infection are in progress.

scholarly journals Topological properties of the triple gene block protein 2 of Bamboo mosaic virus

Virology ◽  
2008 ◽  
Vol 379 (1) ◽  
pp. 1-9 ◽  
Author(s):  
Hsiu-Ting Hsu ◽  
Yuan-Lin Chou ◽  
Yang-Hao Tseng ◽  
Yu-Hsing Lin ◽  
Tzung-Min Lin ◽  
...  
Keyword(s):  
Mosaic Virus ◽  
Triple Gene Block ◽  
Topological Properties ◽  
Gene Block ◽  
Triple Gene Block Protein
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