Varicella-Zoster virus ORF9p: role in viral envelopment and cell-cell fusion

Varicella-Zoster virus is a double-stranded DNA virus belonging to the Herpesviridae family. Its genome encodes 72 ORFs among which ORF9p, the most expressed protein during the lytic cycle of the virus.
Previous work in our laboratory showed that ORF9p plays a role in viral envelopment and that it may interact with the cellular Adaptor Protein-1 (AP-1) complex to do so.
The aim of this work was to better characterize the role of ORF9p during secondary envelopment and to understand the importance of the ORF9p/AP-1 interaction in this process.
First, we confirmed the ORF9p/AP-1 interaction in an infection context and identified the residue leucine 231 of ORF9p as critical for this interaction. We showed that alanine substitution of this residue led to a mislocalization of ORF9p in infected cells as well as a strong replication defect of the virus. Then, we showed that the L231A mutation was also associated with a lack of cell-cell fusion usually seen upon infection by the Wild-type virus. Moreover, naturally occurring compensatory mutations in the coding sequences of gE and/or gH not only restored the fusion phenotype, but also increased cell-cell fusion compared to the Wild-type virus. We have also demonstrated that ORF9p favored the formation of glycoproteins complexes around gE and gH and that the ORF9p L231A mutation destabilized these complexes. Finally, we propose a model explaining the potential role of ORF9p in secondary envelopment of VZV as well as in the cell-cell fusion process.