H-1 parvovirus-associated replication bodies: A distinct virus-induced nuclear structure

We have identified a nuclear structure that is induced after infection with the autonomous parvovirus H-1, Using fluorescence microscopy, we observed that the major nonstructural protein (NS1) of H-1 virus which is essential for viral DNA amplification colocalized with virus-specific DNA sequences a nd sites of ongoing viral DNA replication in distinct nuclear bodies which we designated H-1 parvovirus associated replication bodies (H-l PAR-bodies) . In addition, two cellular proteins were shown to accumulate in H1 PAR-bod ies: (i) the proliferating cell nuclear antigen (PCNA) which is essential f or chromosomal and parvoviral replication and (ii) the NS1-interacting smal l glutamine-rich TPR-containing protein (SGT), suggesting a role for the la tter in parvoviral replication and/or gene expression. Since many DNA virus es target preexisting nuclear structures, known as PML bodies, for viral re plication and gene expression, we have determined the localization of H-1 P AR- and PML-bodies by double-fluorescence labeling and confocal microscopy and found them to be spatially unrelated. Furthermore, H-1 PAR-bodies did n ot colocalize with other prominent nuclear structures such as nucleoli, coi led bodies, and speckled domains. Electron microscopy analysis revealed tha t NS1, as detected by indirect immunogold labeling, was localized in ring-s haped electron-dense nuclear structures corresponding in size and frequency to H-1 PAR-bodies. These structures were also clearly visible without immu nogold labeling and could be detected only in infected cells. Our results s uggest that H-1 virus does not target known nuclear bodies for DNA replicat ion but rather induces the formation of a novel structure in the nucleus of infected cells.