Genome-Wide Screen of Three Herpesviruses for Protein Subcellular Localization and Alteration of PML Nuclear Bodies

Abstract

Herpesviruses are large, ubiquitous DNA viruses with complex host interactions, yet many of the proteins encoded by these viruses have not been functionally characterized. As a first step in functional characterization, we determined the subcellular localization of 234 epitope-tagged proteins from herpes simplex virus, cytomegalovirus, and Epstein–Barr virus. Twenty-four of the 93 proteins with nuclear localization formed subnuclear structures. Twelve of these localized to the nucleolus, and five at least partially localized with promyelocytic leukemia (PML) bodies, which are known to suppress viral lytic infection. In addition, two proteins disrupted Cajal bodies, and 19 of the nuclear proteins significantly decreased the number of PML bodies per cell, including six that were shown to be SUMO-modified. These results have provided the first functional insights into over 120 previously unstudied proteins and suggest that herpesviruses employ multiple strategies for manipulating nuclear bodies that control key cellular processes.

Viral and cellular subnuclear structures in human cytomegalovirus-infected cells

“…These proteins are expressed in different kinetic classes of viral transcription and have different effects on HCMV replication, plus the function of several of these proteins is unknown (Table 1). In transfection experiments epitope-tagged HCMV proteins US32, UL80a and UL35 co-localize with, but do not obviously degrade, PML-NBs in uninfected cells (Salsman et al, 2008). Epitope-tagged US32 and UL35 alter the shape and size of PML-NBs in uninfected cells.…”

“…However, transfection of epitope-tagged HCMV UL30 leads to disruption and/or loss of Cajal bodies in uninfected cells (Table 1). Thus, it has been suggested (Salsman et al, 2008) that UL30 may allow HCMV to inhibit transcription and ribonuclear particle maturation. Cajal bodies are thought to play a role in adenovirus replication, wherein adenovirus infection results in fragmentation of Cajal bodies (James et al, 2010).…”

“…Expression of epitope-tagged versions of US25, UL29 UL30, UL69, UL76, UL98 or TLR9 is sufficient to modestly reduce the number of PML-NBs in uninfected cells (Salsman et al, 2008). These proteins are expressed in different kinetic classes of viral transcription and have different effects on HCMV replication, plus the function of several of these proteins is unknown (Table 1).…”

“…This mechanism of cellular defence to infection has not yet been observed in HCMVinfected cells. An epitope-tagged version of the putative HCMV protein UL3 has been reported to alter PML-NBs in uninfected cells (Salsman et al, 2008). However, it is now thought that the HCMV genome does not contain an ORF encoding UL3 (Gatherer et al, 2011).…”

“…It has been reported that HCMV pp65 and UL84 proteins are a nucleolar antigens (Arcangeletti et al, 2009;Bender et al, 2014). Also, transfection experiments have indicated that several epitopetagged versions of HCMV proteins (TLR5, TLR7, US33, UL29, UL31 and UL76) localize to the nucleolus in uninfected cells (Salsman et al, 2008). These proteins are expressed in different kinetic classes of viral RNA transcription and have different effects on HCMV replication, plus the function of several of these proteins is unknown (Table 1).…”