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Promyelocytic leukemia (PML) nuclear bodies (also known as ND10) are nuclear substructures that contain several proteins, including PML itself, Sp100, and hDaxx. PML has been implicated in many cellular processes, and ND10 are frequently associated with the replicating genomes of DNA viruses. During herpes simplex virus type 1 (HSV-1) infection, the viral regulatory protein ICP0 localizes to ND10 and induces the degradation of PML, thereby disrupting ND10 and dispersing their constituent proteins. ICP0-null mutant viruses are defective in PML degradation and ND10 disruption, and concomitantly they initiate productive infection very inefficiently. Although these data are consistent with a repressive role for PML and/or ND10 during HSV-1 infection, evidence in support of this hypothesis has been inconclusive. By use of short interfering RNA technology, we demonstrate that depletion of PML increases both gene expression and plaque formation by an ICP0-negative HSV-1 mutant, while having no effect on wild-type HSV-1. We conclude that PML contributes to a cellular antiviral repression mechanism that is countered by the activity of ICP0.

Several viruses, including human cytomegalovirus (HCMV), encode proteins that colocalize with a cellular subnuclear structure known as ND10. Since only viral DNA deposited at ND10 initiates transcription, ND10 structures were hypothesized to be essential for viral replication. On the other hand, interferon treatment induces an up-regulation of ND10 structures and viruses have evolved polypeptides that disperse the dot-like accumulation of ND10 proteins, suggesting that ND10 could also be part of an intrinsic defense mechanism. In order to obtain evidence for either a proviral or an antiviral function of ND10, we generated primary human fibroblasts with a stable, short interfering RNA-mediated knockdown (kd) of PML. In these cells, other ND10-associated proteins like hDaxx showed a diffuse nuclear distribution. Interestingly, we observed that HCMV infection induced the de novo formation of ND10-like hDaxx and Sp100 accumulations that colocalized with IE2 and were disrupted, in the apparent absence of PML, in an IE1-dependent manner during the first hours after infection. Furthermore, infection of PML-kd cells with wild-type HCMV at a low multiplicity of infection resulted in enhanced replication. In particular, a significantly increased plaque formation was detected, suggesting that more cells are able to support initiation of replication in the absence of PML. While there was no difference in viral DNA uptake between PML-kd and control cells, we observed a considerable increase in the number of immediate-early (IE) protein-positive cells, indicating that the depletion of PML augments the initiation of viral IE gene expression. These results strongly suggest that PML functions as part of an intrinsic immune mechanism against cytomegalovirus infections.

Research in the last 2 decades has demonstrated that a specific organelle of the cell nucleus, termed PML nuclear body (PML-NB) or nuclear domain 10 (ND10), is frequently modified during viral infection. This correlates with antagonization of a direct repressive function of individual PML-NB components, such as the PML, hDaxx, Sp100, or ATRX protein, that are able to act as cellular restriction factors. Recent studies now reveal an emerging role of PML-NBs as coregulatory structures of both type I and type II interferon responses. This emphasizes that targeting of PML-NBs by viral regulatory proteins has evolved as a strategy to compromise intrinsic antiviral defense and innate immune responses.