Research Projects

Characterization of a host factor that interacts with the Tomato bushy stunt virus (TBSV) cell-to-cell movement protein, P22

Biochemical and biological analyses of TBSV P19

P19-mediated suppression of gene silencing

Analyses of Hin19, a host-encoded RNA processing protein that interacts with P19

Characterization of a novel plant virus

Host-dependent effects on RNA recombination

SPMV Capsid Protein: A Role in Host-Specific Movement.

SPMV as an Epitope Display Vector

The long 3'- untranslated region (UTR) on SPMV RNA.

Transgenic Proso Millet: Characterization of Host-Virus and PMV-SPMV Interactions.

Characterization of a host factor that interacts with the Tomato bushy stunt virus (TBSV) cell-to-cell movement protein, P22 ( representative paper )

This projects aims to analyze the interaction between P22 and a host homeodomain leucine-zipper transcription factor (HFi22), and how this influences infection of plants with TBSV. Recent activities include the detection, localization and sub-cellular distribution of HFi22 in healthy and infected plants. Transgenic plants are being generated to examine the effect of HFi22 over-expression or silencing on plant development and TBSV infection

Biochemical and biological analyses of TBSV P19 (representative paper)

P19 has host-dependent roles in infection and recent emphasis is placed on its function as a suppressor of gene silencing. Within this context, we examine the biological relevance of its biochemical properties to form dimers and to bind siRNAs during infection. For this purpose P19 and mutant versions are compared upon purification from E. coli and infected plants.

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P19-mediated suppression of gene silencing (representative paper)

The suppression function of P19 protects TBSV RNA from gene silencing mediated RNA degradation. We are analyzing amino acid substitution mutants for their ability to protect TBSV RNA from degradation during infection. In addition we are generating transgenic plants expressing non-toxic P19 variants to examine the effect of constitutive suppression on plant development and susceptibility to virus infection.

Analyses of Hin19, a host-encoded RNA processing protein that interacts with P19 (representative paper)

Hin19 is a member of a class of proteins involved in RNA transcription and/or nuclear export, and it specifically interacts with P19. We are presently investigating the localization and sub-cellular distribution of Hin19, in presence or absence of P19. In addition, we are generating transgenic plants that over-express Hin19 or those that are silenced for its expression and to test these for virus infection and the ability of P19 to suppress gene silencing.

Characterization of a novel plant virus (representative paper)

This project aims to characterize a new plant virus that infects maize and wheat. Experiments thus far show that the virus is transmitted by the wheat curl mite and enveloped particles encapsidate an RNP complex that contains 4-5 RNA species of negative or ambisense polarity. Sequence analyses do not show any similarity with any known viral agents. The virus interacts in novel ways with Wheat streak mosaic virus during mixed infections

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Host-dependent effects on RNA recombination (representative papers: One, Two)

The two above referenced studies each provided independent evidence that virus RNA recombination is host-dependent. Experiments are planned to further examine this phenomenon by studying the underlying mechanism and at which level the host-dependent prevention or stimulation of RNA recombination occurs.

SPMV Capsid Protein: A Role in Host-Specific Movement.

Recently, we determined that the 17-kDa SPMV capsid protein (CP) contributes to and/or coordinates long distance movement in foxtail millet. SPMV is an 824 nucleotide (nt) RNA that is completely dependent on Panicum mosaic virus for replication in millet plants. Subcellular fractionation has revealed that the SPMV CP accumulates in the cytosol, but it is primarily concentrated in cell wall-enriched fractions, a hallmark of plant virus movement proteins. Moreover, we have found that the C-terminal region of the SPMV CP is specifically associated with membrane fractions and this same region is responsible for a severe symptom phenotype on millet plants.

SPMV as an Epitope Display Vector.

The intent of this project is to develop novel novel veterinary vaccines for the protection of our food supply. Plant viruses are routinely consumed by both animals and humans. Thus far, they have not been reported to cause negative health effects such as allergic responses. Therefore, our intent is to produce use satellite panicum mosaic virus (SPMV) an an epitope display vector. This may provide a new production and delivery system for cost-effective bulk production of vaccines in grasses. The influenza A virus hemagglutinin epitope (HA.11) has been inserted into five exposed regions on the SPMV capsid protein, based on the known x-ray crystal structure. Our current research is directed toward determining if the HA.11 mutants are a) viable when inoculated on millet plants, b) immunogenic in poultry, and c) can be successfully deployed for injection and/or oral-mucosal primed immune responses.

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The long 3'- untranslated region (UTR) on SPMV RNA.

In our previous studies we have observed that the entire 3'-UTR is necessary for replication of SPMV in millet plants. This suggests that it has a crucial role in ensuring the interaction with the PMV replicase in trans, and possible is associated with RNA movement. From this, we have designed several deletion and insertion mutants to evaluate the importance of this sequence in SPMV biology, to include encapsidation and translation of the SPMV CP gene.

Transgenic Proso Millet: Characterization of Host-Virus and PMV-SPMV Interactions.

Thus far millet plants have been intransigent to Agrobacterium-mediated transformation, although it is possible to regenerate both proso and foxtail millet. To further our understanding of the role of the PMV replicase proteins in supporting SPMV in proso millet we are exploring several strategies to produce transgenic plants. In addition, SPMV CP and various truncations of the CP are also of interest to further evaluate cellular localization and host responses to this