A Conserved Helix in the C-Terminal Region of Watermelon Silver Mottle Virus Nonstructural Protein S Is Imperative For Protein Stability Affecting Self-Interaction, RNA Silencing Suppression, and Pathogenicity
| Autor: | Joseph A. J. Raja, Shih-Shun Lin, Tzu-Tung Lin, Chung-Hao Huang, Yue-Rong Tan, Shyi-Dong Yeh, Mung-Hsia Foo |
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| Rok vydání: | 2020 |
| Předmět: |
0106 biological sciences
Small interfering RNA Physiology Nicotiana benthamiana Viral Nonstructural Proteins medicine.disease_cause 01 natural sciences Protein S 03 medical and health sciences Bimolecular fluorescence complementation Tospovirus RNA interference Tobacco medicine Turnip mosaic virus 030304 developmental biology chemistry.chemical_classification 0303 health sciences Mutation Virulence biology Protein Stability General Medicine biology.organism_classification Cell biology Amino acid RNA silencing chemistry RNA Interference Agronomy and Crop Science 010606 plant biology & botany |
| Zdroj: | Molecular Plant-Microbe Interactions®. 33:637-652 |
| ISSN: | 1943-7706 0894-0282 |
| Popis: | In orthotospovirus, the nonstructural protein S (NSs) is the RNA-silencing suppressor (RSS) and pathogenicity determinant. Here, we demonstrate that a putative α-helix, designated H8, spanning amino acids 338 to 369 of the C-terminal region of the NSs protein, is crucial for self-interaction of watermelon silver mottle virus NSs protein and that the H8 affects RSS function. Co-immunoprecipitation, yeast two-hybrid, and bimolecular fluorescence complementation analyses revealed that the triple point mutation (TPM) of H8 amino acids Y338A, H350A, and F353A resulted in NSs protein self-interaction dysfunction. Transient expression of H8-deleted (ΔH8) and TPM NSs proteins in Nicotiana benthamiana plants by agroinfitration indicated that these proteins have weaker RSS activity and are far less stable than wild-type (WT) NSs. However, an electrophoretic mobility assay revealed that small interfering RNA (siRNA) binding ability of TPM NSs protein is not compromised. The pathogenicity assay of WT NSs protein expressed by the attenuated turnip mosaic virus vector restored severe symptoms in recombinant-infected N. benthamiana plants but not for ΔH8 or TPM proteins. Taken together, we conclude that the H8 helix in the C-terminal region of NSs protein is crucial for stabilizing NSs protein through self-interaction to maintain normal functions of RSS and pathogenicity, but not for NSs-siRNA binding activity. |
| Databáze: | OpenAIRE |
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