Ser/Thr Kinase-Like Protein of Nicotiana benthamiana Is Involved in the Cell-to-Cell Movement of Bamboo mosaic virus

Cheng, Shun-Fang; Tsai, Ming Chieh; Huang, Chia-Lin; Huang, Y. S.; Chen, I‐Hsuan; Lin, Na-Sheng; Hsu, Yau‐Heiu; Tsai, Ching‐Hsiu; Cheng, Chi-Ping

doi:10.1371/journal.pone.0062907

Cited by 21 publications

(15 citation statements)

References 40 publications

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“…For VIGS, the TRV‐based knockdown system was used, as described previously (Cheng et al ., ). Briefly, agrobacteria containing plasmid TRV1 and TRV2 derivatives were mixed in a 1 : 1 ratio and infiltrated into the second, third and fourth true leaves of five‐leaf stage N. benthamiana .…”

Section: Methodsmentioning

confidence: 97%

“…The procedure of transient expression has been described previously (Cheng et al ., ). In general, after induction with acetosyringone, agrobacteria containing NbRABG3f or its derivatives were mixed with Agrobacterium harbouring the silencing suppressor HcPro (Chiu et al ., ) in a 1 : 1 ratio and infiltrated onto the third, fourth and fifth leaves of N. benthamiana at the six‐leaf stage.…”

Section: Methodsmentioning

confidence: 97%

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NbRABG3f, a member of Rab GTPase, is involved in Bamboo mosaic virus infection in Nicotiana benthamiana

Huang

Jhuo

Tsai

et al. 2015

Molecular Plant Pathology

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The screening of differentially expressed genes in plants after pathogen infection can uncover the potential host factors required for the pathogens. In this study, an up-regulated gene was identified and cloned from Nicotiana benthamiana plants after Bamboo mosaic virus (BaMV) inoculation. The up-regulated gene was identified as a member of the Rab small guanosine triphosphatase (GTPase) family, and was designated as NbRABG3f according to its in silico translated product with high identity to that of RABG3f of tomato. Knocking down the expression of NbRABG3f using a virus-induced gene silencing technique in a protoplast inoculation assay significantly reduced the accumulation of BaMV. A transiently expressed NbRABG3f protein in N. benthamiana plants followed by BaMV inoculation enhanced the accumulation of BaMV to approximately 150%. Mutants that had the catalytic site mutation (NbRABG3f/T22N) or had lost their membrane-targeting capability (NbRABG3f/ΔC3) failed to facilitate the accumulation of BaMV in plants. Because the Rab GTPase is responsible for vesicle trafficking between organelles, a mutant with a fixed guanosine diphosphate form was used to identify the donor compartment. The use of green fluorescent protein (GFP) fusion revealed that GFP-NbRABG3f/T22N clearly co-localized with the Golgi marker. In conclusion, BaMV may use NbRABG3f to form vesicles derived from the Golgi membrane for intracellular trafficking to deliver unidentified factors to its replication site; thus, both GTPase activity and membrane-targeting ability are crucial for BaMV accumulation at the cell level.

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Section: Methodsmentioning

confidence: 97%

Section: Methodsmentioning

confidence: 97%

NbRABG3f, a member of Rab GTPase, is involved in Bamboo mosaic virus infection in Nicotiana benthamiana

Huang

Jhuo

Tsai

et al. 2015

Molecular Plant Pathology

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“…The image was taken at 3 days post-infiltration of the leaf tissue or the protoplasts with a confocal laser scanning microscope (FV1000, Olympus) using a 403 objective lens. For the fractionation assay, the soluble and membrane fractions were separated as described previously (Cheng et al, 2013b).…”

Section: Localization Of Nbtrxh2 and Its Derivativesmentioning

confidence: 99%

A thioredoxin NbTRXh2 from Nicotiana benthamiana negatively regulates the movement of Bamboo mosaic virus

Chen

Huang

et al. 2017

Molecular Plant Pathology

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An up-regulated gene derived from Bamboo mosaic virus (BaMV)-infected Nicotiana benthamiana plants was cloned and characterized in this study. BaMV is a single-stranded, positive-sense RNA virus. This gene product, designated as NbTRXh2, was matched with sequences of thioredoxin h proteins, a group of small proteins with a conserved active-site motif WCXPC conferring disulfide reductase activity. To examine how NbTRXh2 is involved in the infection cycle of BaMV, we used the virus-induced gene silencing technique to knock down NbTRXh2 expression in N. benthamiana and inoculated the plants with BaMV. We observed that, compared with control plants, BaMV coat protein accumulation increased in knockdown plants at 5 days post-inoculation (dpi). Furthermore, BaMV coat protein accumulation did not differ significantly between NbTRXh2-knockdown and control protoplasts at 24 hpi. The BaMV infection foci in NbTRXh2-knockdown plants were larger than those in control plants. In addition, BaMV coat protein accumulation decreased when NbTRXh2 was transiently expressed in plants. These results suggest that NbTRXh2 plays a role in restricting BaMV accumulation. Moreover, confocal microscopy results showed that NbTRXh2-OFP (NbTRXh2 fused with orange fluorescent protein) localized at the plasma membrane, similar to AtTRXh9, a homologue in Arabidopsis. The expression of the mutant that did not target the substrates failed to reduce BaMV accumulation. Co-immunoprecipitation experiments revealed that the viral movement protein TGBp2 could be the target of NbTRXh2. Overall, the functional role of NbTRXh2 in reducing the disulfide bonds of targeting factors, encoded either by the host or virus (TGBp2), is crucial in restricting BaMV movement.

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“…Of those factors, a glutathione transferase (NbGSTU4) promotes BaMV accumulation presumably by providing a more suitable redox environment for BaMV replication ( Chen et al, 2013 ). A serine/threonine kinase-like protein (NbSTKL) is involved in the cell-to-cell movement of BaMV ( Cheng et al, 2013b ), while a putative Rab-GTPase activation protein (NbRabGAP1) is important for the viral intercellular movement ( Huang et al, 2013 ).…”

Section: Introductionmentioning

confidence: 99%

Promotion of Bamboo Mosaic Virus Accumulation in Nicotiana benthamiana by 5′→3′ Exonuclease NbXRN4

Lee

Lin

et al. 2016

Front. Microbiol.

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Bamboo mosaic virus (BaMV) has a 6.4-kb (+) sense RNA genome with a 5′ cap and a 3′ poly(A) tail. ORF1 of this potexvirus encodes a 155-kDa replication protein responsible for the viral RNA replication/transcription and 5′ cap formation. To learn more about the replication complex of BaMV, a protein preparation enriched in the 155-kDa replication protein was obtained from Nicotiana benthamiana by a protocol involving agroinfiltration and immunoprecipitation. Subsequent analysis by SDS-PAGE and mass spectrometry identified a handful of host proteins that may participate in the viral replication. Among them, the cytoplasmic exoribonuclease NbXRN4 particularly caught our attention. NbXRN4 has been shown to have an antiviral activity against Tomato bushy stunt virus and Tomato mosaic virus. In Arabidopsis, the enzyme could reduce RNAi- and miRNA-mediated RNA decay. This study found that downregulation of NbXRN4 greatly decreased BaMV accumulation, while overexpression of NbXRN4 resulted in an opposite effect. Mutations at the catalytically essential residues abolished the function of NbXRN4 in the increase of BaMV accumulation. Nonetheless, NbXRN4 was still able to promote BaMV accumulation in the presence of the RNA silencing suppressor P19. In summary, the replication efficiency of BaMV may be improved by the exoribonuclease activity of NbXRN4.

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Ser/Thr Kinase-Like Protein of Nicotiana benthamiana Is Involved in the Cell-to-Cell Movement of Bamboo mosaic virus

Cited by 21 publications

References 40 publications

NbRABG3f, a member of Rab GTPase, is involved in Bamboo mosaic virus infection in Nicotiana benthamiana

NbRABG3f, a member of Rab GTPase, is involved in Bamboo mosaic virus infection in Nicotiana benthamiana

A thioredoxin NbTRXh2 from Nicotiana benthamiana negatively regulates the movement of Bamboo mosaic virus

Promotion of Bamboo Mosaic Virus Accumulation in Nicotiana benthamiana by 5′→3′ Exonuclease NbXRN4

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