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Functional Characterization of Suppressors of Gene Silencing and Development of an A. thaliana Transgenic Plant to Study the TSWV Silencing Suppressor NSS Protein

Rosalba Rodriguez-Pena, University of Nebraska - Lincoln

Abstract

Antiviral gene silencing is a plant defense mechanism that involves Dicer-like proteins, small interfering RNAs (siRNA), Argonautes (AGO), RNA-dependent RNA-polymerases (RDR), and Suppressor of gene silencing co-factor 3 (SGS3). Viruses encode viral suppressor of RNA silencing (VSR) to overcome the antiviral silencing. Sugarcane mosaic virus (SCMV), Maize chlorotic mottle virus (MCMV), and Tomato spotted wilt virus (TSWV) are important plant viruses. SCMV VPg and MCMV P50 proteins were suggested to be the VSR, however, the mechanism of suppression is unknown or unclear. In the case of TSWV, the Non-structural small protein (NSs) is the VSR, that bind both long and small double-stranded RNA. Nevertheless, the suppression mechanism is not well-understood. Here, we analyzed the pattern of relocalized plant proteins induced by virus infection through a review of research articles. The review showed that host proteins are relocalized mainly from the cytoplasm to the viruses’ replication compartments, and the relocalizations have a proviral effect on viruses’ replication. We also co-expressed VPg and P50 individually with AGO1, AGO2, AGO4, AGO5, AGO7, AGO10, RDR1, RDR6, and SGS3. We found that SCMV VPg targets AGO2, RDR6, and SGS3 for degradation, and RDR6 degradation is mediated via 20 S proteosome. The exact mechanism of the MCMV P50 suppression remained unsolved. Interesting, the upregulation of all tested plant proteins was observed. This result suggests that MCMV P50 may target siRNA for sequestration and/or disrupt systemic movement. To study the NSs suppression mechanism we developed an Arabidopsis thaliana transgenic plant by the floral dipped transformation method. The insertion of NSs in the plant genome and the expression of the transcript were detected, however, we were unable to detect the protein. These results suggested that the NSs protein may be present at a very low concentration, or the mRNA is untranslatable. The research advances the understating of the host-viral protein interaction during the antiviral silencing pathway and suppression mechanism.

Subject Area

Virology|Plant Pathology|Plant sciences

Recommended Citation

Rodriguez-Pena, Rosalba, "Functional Characterization of Suppressors of Gene Silencing and Development of an A. thaliana Transgenic Plant to Study the TSWV Silencing Suppressor NSS Protein" (2022). ETD collection for University of Nebraska - Lincoln. AAI29259200.
https://digitalcommons.unl.edu/dissertations/AAI29259200

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