Visualizing tomato spotted wilt virus protein localization: Cross-kingdom comparisons of protein-protein interactions.

Tomato spotted wilt virus (TSWV) is an orthotospovirus that infects both plant and insect cells. Understanding the protein localization and interactions in these cells is crucial for unraveling the infection cycle and host-virus interactions. In this study, we investigated the localization of TSWV proteins in cells of plants and insects. Furthermore, we identified the protein-protein interactions among TSWV proteins using bimolecular fluorescence complementation (BiFC) and yeast two-hybrid (MbY2H) assays. Our results revealed distinct localization patterns for TSWV proteins in plant and insect cells. The nucleocapsid protein (N), essential for genome encapsidation, was found in the cytoplasm of both cell types. The non-structural movement protein (NSm) localized to the cytoplasm in insect cells, different from the localization in plant cells plasmodesmata. The non-structural silencing protein (NSs) exhibited peripheral localization in plant cells and cytoplasmic localization in insect cells. Additionally, glycoproteins GN and GC showed cytoplasmic localization in both cell types. Moreover, protein-protein interaction analyses revealed self-interactions of NSm, N, GN, GC, and NSs. These interactions are crucial for viral genome encapsidation, virion assembly, and RNA silencing suppression. We also identified interactions between different TSWV proteins, indicating their roles and host interactions. Comparisons with other orthotospovirus interaction maps highlighted the uniqueness of TSWV protein-protein interaction networks. Despite sharing genome organization and putative gene annotations, each orthotospovirus exhibited distinct interaction maps. Overall, this research expands our knowledge of TSWV infection and elaborates on the intricate relationships between viral proteins, cellular dynamics, and host responses. These findings lay the groundwork for future studies on the molecular mechanisms of TSWV infection and may facilitate the development of effective control strategies.