Human cytomegalovirus antagonizes SMC5/6 driven genome silencing via UL35 instituted proteasomal degradation of SLF2

Recent studies demonstrated that the structural maintenance of chromosomes complex 5/6 (SMC5/6) acts as an important antiviral restriction factor that silences episomal DNA. Consequently, viruses have evolved strategies to antagonize this defence system. Interestingly, it was shown that SMC5/6-mediated silencing of extrachromosomal DNA depends on its recruitment to PML nuclear bodies which is mediated by a protein termed SMC5/6 localization factor 2 (SLF2). Since human cytomegalovirus (HCMV) dissociates PML nuclear bodies (PML-NBs) during the first hours of infection, we asked for the fate of SMC5/6 components during the HCMV replicative cycle. We first investigated the expression levels of SMC5/6 core components at the onset of HCMV infection and found that they were not downregulated by HCMV. Instead, we observed a distinct decrease of SLF2 protein levels which correlated with a complete dispersal of the SMC5/6 complex from PML-NBs. This was also observed after infection with UV inactivated HCMV and could not be blocked by cycloheximide treatment suggesting the involvement of an imported structural component of the virion. While we could exclude a role of the HCMV tegument protein pp71 (UL82), a targeted screen identified the tegument protein UL35 as being required and sufficient for proteasomal degradation of SLF2. In line with these results, SLF2 levels remained stable after infection with a recombinant HCMV harbouring a stop codon within the UL35 gene region and this correlated with a defect in the onset of HCMV immediate early gene expression. Furthermore, we observed an enhanced recruitment of SMC5/6 to PML-NBs and an overall increase in the association of parental viral genomes with SMC5/6 containing PML-NBs in the absence of UL35. Congruently, siRNA mediated depletion of SLF2 resulted in a reversal of this phenotype. This strongly suggests that UL35 serves as a novel viral SLF2 antagonist to prevent the initial silencing of incoming viral genomes by SMC5/6. Author summaryHuman cytomegalovirus (HCMV) is an important human pathogen which establishes a life-long persistent infection. Previous research revealed that host intrinsic defense mechanisms can silence HCMV gene expression and thereby foster latency. However, during lytic infection, viral effector proteins like the structural protein pp71 (UL82) and the viral immediate-early protein IE1 act as antagonists of this defense. Here, we report on the identification of a novel antagonist of host-mediated viral gene silencing which targets the SMC5/6 complex. SMC5/6 belongs to a highly conserved family of protein complexes that act as loop extrusion machines to facilitate the folding of chromatin. While SMC5/6 core factors were not attacked by HCMV, we found that the SMC5/6 associated protein SLF2 was degraded by the viral tegument protein UL35, that is imported into infected cells. This abrogates the accumulation of SMC5/6 at PML nuclear bodies which compromises the entrapment of viral genomes in these subnuclear structures. Since SLF2 is also important for genome stability and pathogenic variants of SLF2 cause the neurodevelopmental disease Atelis syndrome, we speculate that depletion of this factor via a viral tegument protein may also contribute to DNA damage induced during congenital HCMV infection.