NSP2 forms viroplasms during Dendrolimus punctatus cypovirus infection

Reoviruses are thought to replicate and assemble in special cytoplasmic structures called 'viroplasms'. However, little is known about the viroplasms of the insect reoviruses, the cypoviruses. To investigate the viroplasm of Dendrolimus punctatus cypovirus (DpCPV), all proteins encoded by the 10 genomic segments of DpCPV were expressed in Sf9 cells using the Bac-to-Bac system. The viral nonstructural protein NSP2 formed viroplasm-like dots which showed close apposition with the endoplasmic reticulum and were surrounded by intracellular membranes during transfection. Colocalization and coimmunoprecipitation assays showed that NSP2 interacts with 4 of 6 structural proteins and another 2 nonstructural proteins, while NSP1 only colocalized with VP4, and NSP3 did not colocalize with any structural protein. Immunoelectron microscopy revealed that NSP2 were nearby the endoplasmic reticulum and mitochondria, and viral particles were present in the electron-dense inclusions formed by NSP2. We proposed that NSP2 is responsible for forming the viroplasms structures of DpCPV.