Role of Kinases in VZV Nuclear Egress

Like all Herpesviruses VZV expresses two conserved proteins forming the nuclear egress complex (NEC), namely ORF24p and ORF27p, respectively homologous to UL34 and UL31 in HSV-1. Many works described the role of this complex and viral kinases, the Alpha-herpesvirus specific kinase US3p, and hCMV UL97p (conserved in all Herpesviruses and homolog to UL13p in HSV-1), in the nuclear capsids translocation to the cytoplasm. Indeed, using a very complex mechanism, not yet fully characterized, nuclear capsids bud through the internal nuclear membrane, stay briefly in the perinuclear space and finally free the capsids into the cytoplasm through fusion of the primary envelope with the external nuclear membrane. In the cytoplasm, virions acquire the second and last envelopment before being released outside the cells. In order to better characterize the nuclear egress in the context of VZV, we focused our work on the NEC and on both viral kinases ORF47p and ORF66p respectively homologous to HSV-1 UL13p and US3p. By co-immunoprecipitation we identified a protein complex containing ORF24p, ORF27p and both viral kinases and are currently trying to determine the role of each kinase in the formation of this complex. We are also characterizing the phosphorylation status of these proteins depending on ORF66p and/or ORF47p. Immunofluorescence studies of cells infected with a deleted ORF66 or ORF47 virus showed mislocalisation of some viral proteins such as ORF24p as described for HSV-1. Finally electron microscopy analyses are in progress and will help to determine the role of both VZV kinases in the nuclear egress.