Identification of TRIM27 as a Novel Degradation Target of Herpes Simplex Virus 1 ICP0

Abstract

The herpes simplex virus 1 (HSV-1) immediate early protein ICP0 performs many functions during infection, including transactivation of viral gene expression, suppression of innate immune responses, and modification and eviction of histones from viral chromatin. Although these functions of ICP0 have been characterized, the detailed mechanisms underlying ICP0's complex role during infection warrant further investigation. We thus undertook an unbiased proteomic approach to identifying viral and cellular proteins that interact with ICP0 in the infected cell. Cellular candidates resulting from our analysis included the ubiquitinspecific protease USP7, the transcriptional repressor TRIM27, DNA repair proteins NBN and MRE11A, regulators of apoptosis, including BIRC6, and the proteasome. We also identified two HSV-1 early proteins involved in nucleotide metabolism, U L 39 and U L 50, as novel candidate interactors of ICP0. Because TRIM27 was the most statistically significant cellular candidate, we investigated the relationship between TRIM27 and ICP0. We observed rapid, ICP0-dependent loss of TRIM27 during HSV-1 infection. TRIM27 protein levels were restored by disrupting the RING domain of ICP0 or by inhibiting the proteasome, arguing that TRIM27 is a novel degradation target of ICP0. A mutant ICP0 lacking E3 ligase activity interacted with endogenous TRIM27 during infection as demonstrated by reciprocal coimmunoprecipitation and supported by immunofluorescence data. Surprisingly, ICP0-null mutant virus yields decreased upon TRIM27 depletion, arguing that TRIM27 has a positive effect on infection despite being targeted for degradation. These results illustrate a complex interaction between TRIM27 and viral infection with potential positive or negative effects of TRIM27 on HSV under different infection conditions. IMPORTANCEDuring productive infection, a virus must simultaneously redirect multiple cellular pathways to replicate itself while evading detection by the host's defenses. To orchestrate such complex regulation, viruses, including herpes simplex virus 1 (HSV-1), rely on multifunctional proteins such as the E3 ubiquitin ligase ICP0. This protein regulates various cellular pathways concurrently by targeting a diverse set of cellular factors for degradation. While some of these targets have been previously identified and characterized, we undertook a proteomic screen to identify additional targets of this activity to further characterize ICP0's role during infection. We describe a set of candidate interacting proteins of ICP0 identified through this approach and our characterization of the most statistically significant result, the cellular transcriptional repressor TRIM27. We present TRIM27 as a novel degradation target of ICP0 and describe the relationship of these two proteins during infection. During lytic infection, a virus must hijack the synthesis machinery of its host cell to manufacture its own components. The virus redirects cellular metabolism, chromatin regulation, transcription factors, and t...

Time-resolved Global and Chromatin Proteomics during Herpes Simplex Virus Type 1 (HSV-1) Infection

“…We then sorted for proteins present only in the first three conditions (mock, 3 and 6 hpi). Interestingly, ϳ50% of these were involved in immune response processes (supplemental Table S1C) known to be suppressed upon virus infection (35). Several of these proteins are reported to be degraded upon viral infection such as ITCH, which was found downregulated at 9-15 hpi (36).…”

MG132 exerts anti-viral activity against HSV-1 by overcoming virus-mediated suppression of the ERK signaling pathway

“…The cytoplasmic ICP0 inhibited IRF3 activation and IRF3-dependent ISG induction at later stages of infection. Moreover, proteolytic activities of the proteasome during the HSV-1 infection have been known to be involved in various viral events including inactivation of antiviral response such as degradation of the tegument protein VP11/12 (UL46) in an ICP0-dependent manner 43 , degradation of the transcriptional repressor TRIM27 in an ICP0-dependent manner 44 , degradation of the interferon-inducible protein X (IFIX) recognized as an antiviral factor 45 and degradation of TANK binding kinase 1 (TBK1) in a Us11 (dsRNA-dependent protein kinase/PKR viral inhibitory protein)-dependent mannre 46 . These HSV-1-induced proteasomal degradation phenomena were blocked by MG132 treatment.…”

Cellular Transcriptional Coactivator RanBP10 and Herpes Simplex Virus 1 ICP0 Interact and Synergistically Promote Viral Gene Expression and Replication

“…These experiments identified 80 cell proteins that coimmunoprecipitated with transiently expressed MEF-tagged ICP0 (data not shown). The proteins identified included USP7, which has previously been shown to bind ICP0 (17,46). Of the putative ICP0-interacting cell proteins identified, we focused on RanBP10.…”

“…In contrast, ICP0 also appears to degrade potential positive cellular factors for HSV-1 replication, such as USP7 and TRIM27. USP7 and TRIM27 have been shown to be degraded in HSV-1-infected cells in an ICP0-dependent manner, but both proteins can promote viral replication (14,17,19). (iii) ICP0 has been shown to promote acetylation and eviction of his-tones to modulate the chromatin structure of viral genomes for efficient viral gene expression (20).…”

“…(ii) ICP0 has also been shown to degrade many other cellular proteins, including DNA-dependent protein kinase (DNA-PKcs), RNF8, RNF168, USP7, E2FBP1, IFI16, and TRIM27 (11)(12)(13)(14)(15)(16)(17). It appears that ICP0 degradation of DNA damage regulators DNAPKcs, RNF8, and RNF168 and of IFI16, a sensor of herpesvirus DNAs, counteracts host responses activated by HSV-1 infection, including the DNA damage response and innate immune signaling, respectively (11-13, 16, 18).…”