Protein-protein interactions among common virion structural proteins of Heliothis virescens ascovirus 3h

Ascoviruses are large double-stranded DNA viruses that mainly infect lepidopteran larvae, and may have potential for development as bioinsecticides. Compared with well-studied baculoviruses, interactions among ascovirus virion structural proteins remain unknown. The protein composition of virions of Heliothis virescens ascovirus 3h (HvAV-3h), an ascovirus isolate reported in China, was analyzed using mass spectrometry (MS) in our previous study; however, how the 29 common virion structural proteins interact with one another remained unknown. The primary aim of this study was to systematically identify protein-protein interactions among the 29 common virion structural proteins of HvAV-3h, and further explore the potential assembly pattern of the virion. These proteins were expressed using the Gal4 yeast two-hybrid (Y2H) system. A total of 841 (29 × 29) direct cross Y2H assays were performed, and 192 interactions were identified, including 126 non-self pairwise interactions and 14 self-associations, with the remaining interactions described in the text. The self-associations of 3H-43, 3H-53, and 3H-107 were further confirmed using pull-down assays. A theoretical interaction network of the 29 proteins was constructed using R software. Combined with previously obtained MS data, the possible arrangement patterns of these proteins in HvAV-3h virions produced in Mythimna separata, Spodoptera exigua, Spodoptera frugiperda, and Spodoptera litura larvae were predicted and compared. The results of this study provided a theoretical foundation for understanding ascovirus virion structure and differences in ascovirus pathogenicity among host larval species.