Development of Tomato bushy stunt virus-based vectors for fusion and non-fusion

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Development of Tomato bushy stunt virus-based vectors for fusion and non-fusion expression of heterologous proteins in an alternative host Nicotiana excelsiana 

Plant virus-based expression systems are an alternative expression platform for the production of clinically and industrially useful recombinant proteins. Nonetheless, due to a lack of viral vector with the commercial potentials, it is urgent to design and develop new, versatile, and efficient plant virus vectors.

The genome of Tomato bushy stunt virus (TBSV) offers an attractive alternative to being modified as a vector for producing heterologous proteins in plants.

Here, we developed a set of novel fusion and non-fusion TBSV-CP replacement vectors, which provide more flexible and efficient tools for expressing proteins of interest in plants. An alternative tobacco plant, Nicotiana excelsiana, was used in this study as a host for newly constructed TBSV vectors because the unwanted necrotic effects were reported on the commonly used Nicotiana benthamiana host associated with expression of TBSV-encoded P19 protein.

The data showed that TBSV vectors caused a symptomless infection and overexpressed reporter gene in N. excelsiana leaves, demonstrating that N. excelsiana is an ideal host plant for TBSV-mediated heterologous gene expression. Moreover, a TBSV non-fusion vector, dAUG, shows the similar accumulation level of reporter proteins to that of TMV- and PVX-based vectors in side-by-side comparison and provides more flexible aspects than the previously developed TBSV vectors. Collectively, our newly developed TBSV expression system adds a new member to the family of plant viral expression vectors and meanwhile offers a flexible and highly effective approach for producing proteins of interest in plants.

 

KEY POINTS:

• The TBSV-based transient expression system has been significantly improved.

  • The necrotic effects caused by viral P19 protein were avoided by the usage of N.   excelsiana as a host plant.
  • The expression level of the non-fusion vector was similar to the most effective virus   vectors reported so far.
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Description: Description of target: The protein encoded by this gene is a member of the scavenger receptor cysteine-rich (SRCR) superfamily, and is exclusively expressed in monocytes and macrophages. It functions as an acute phase-regulated receptor involved in the clearance and endocytosis of hemoglobin/haptoglobin complexes by macrophages, and may thereby protect tissues from free hemoglobin-mediated oxidative damage. This protein may also function as an innate immune sensor for bacteria and inducer of local inflammation. Alternatively spliced transcript variants encoding different isoforms have been described for this gene.;Species reactivity: Human;Application: ELISA;Assay info: Assay Methodology: Quantitative Sandwich ELISA;Sensitivity: 0.273 ng/mL

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Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

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116301G0-100tests 100 tests
EUR 547
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

FITC Anti-human CD163 Antibody *GHI/61*

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EUR 2093
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

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116301G1-500tests 500 tests
EUR 2051
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

TRITC Anti-human CD163 Antibody *GHI/61*

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EUR 558
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

TRITC Anti-human CD163 Antibody *GHI/61*

116301H0-100tests 100 tests
EUR 547
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

TRITC Anti-human CD163 Antibody *GHI/61*

116301H1 500 tests
EUR 2093
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

TRITC Anti-human CD163 Antibody *GHI/61*

116301H1-500tests 500 tests
EUR 2051
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

PerCP Anti-human CD163 Antibody *GHI/61*

116301S0 25 tests
EUR 391
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

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EUR 383
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

PerCP Anti-human CD163 Antibody *GHI/61*

116301S1 100 tests
EUR 1015
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

PerCP Anti-human CD163 Antibody *GHI/61*

116301S1-100tests 100 tests
EUR 995
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

PerCP Anti-human CD163 Antibody *GHI/61*

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EUR 3044
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

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EUR 2984
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

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PacBlue Anti-human CD163 Antibody *GHI/61*

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EUR 558
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

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116301I0-100tests 100 tests
EUR 547
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

PacBlue Anti-human CD163 Antibody *GHI/61*

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EUR 2093
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

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EUR 2051
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

Purified Anti-human CD163 Antibody *GHI/61*

11630000 100 ug
EUR 373
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

Purified Anti-human CD163 Antibody *GHI/61*

11630000-100ug 100 ug
EUR 365
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

PacOrange Anti-human CD163 Antibody *GHI/61*

116301J0 100 tests
EUR 558
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

PacOrange Anti-human CD163 Antibody *GHI/61*

116301J0-100tests 100 tests
EUR 547
Description: GHI/61 is an anti-human monoclonal antibody that forms an immune complex with the CD163 antigen.

Effect of Serotype and Strain Diversity on Dengue Virus Replication in Australian Mosquito Vectors

Dengue virus (DENV) is the most important mosquito-borne viral pathogen of humans, comprising four serotypes (DENV-1 to -4) with a myriad of genotypes and strains. The kinetics of DENV replication within the mosquito following ingestion of a blood meal influence the pathogen’s ability to reach the salivary glands and thus the transmission potential. The influence of DENV serotype and strain diversity on virus kinetics in the two main vector species, Aedes aegypti and Ae. albopictus, has been poorly characterized. We tested whether DENV replication kinetics vary systematically among serotypes and strains, using Australian strains of the two vectors.

Mosquitoes were blood fed with two strains per serotype, and sampled at 3, 6, 10 and 14-days post-exposure. Virus infection in mosquito bodies, and dissemination of virus to legs and wings, was detected using qRT-PCR. For both vectors, we found significant differences among serotypes in proportions of mosquitoes infected, with higher numbers for DENV-1 and -2 versus other serotypes.

Consistent with this, we observed that DENV-1 and -2 generally replicated to higher RNA levels than other serotypes, particularly at earlier time points. There were no significant differences in either speed of infection or dissemination between the mosquito species. Our results suggest that DENV diversity may have important epidemiological consequences by influencing virus kinetics in mosquito vectors.

A Plant Virus Ensures Viral Stability in the Hemolymph of Vector Insects through Suppressing Prophenoloxidase Activation

 

Most plant viruses require vector insects for transmission. Viral stability in the hemolymph of vector insects is a prerequisite for successful transmission of persistent plant viruses. However, knowledge of whether the proteolytic activation of prophenoloxidase (PPO) affects the stability of persistent plant viruses remains elusive.

Here, we explored the interplay between rice stripe virus (RSV) and the PPO cascade of the vector small brown planthopper. Phenoloxidase (PO) activity was suppressed by RSV by approximately 60%.

When the PPO cascade was activated, we found distinct melanization around RSV particles and serious damage to viral stability in the hemolymph. Viral suppression of PO activity was derived from obstruction of proteolytic cleavage of PPOs by binding of the viral nonstructural protein NS3. These results indicate that RSV attenuates the PPO response to ensure viral stability in the hemolymph of vector insects. Our research provides enlightening cues for controlling the transmission of vector-borne viruses.

IMPORTANCE Large ratios of vector-borne plant viruses circulate in the hemolymph of their vector insects before entering the salivary glands to be transmitted to plants. The stability of virions in the hemolymph is vital in this process. Activation of the proteolytic prophenoloxidase (PPO) to produce active phenoloxidase (PO) is one of the major innate immune pathways in insect hemolymph.

How a plant virus copes with the PPO immune reaction in its vector insect remains unclear. Here, we report that the PPO affects the stability of rice stripe virus (RSV), a notorious rice virus, in the hemolymph of a vector insect, the small brown planthopper. RSV suppresses PPO activation using viral nonstructural protein. Once the level of PO activity is elevated, RSV is melanized and eliminated from the hemolymph. Our work gives valuable clues for developing novel strategies for controlling the transmission of vector-borne plant viruses.

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