Assembly of Helper-Dependent Adenovirus DNA into Chromatin Promotes Efficient Gene Expression

In infected cells, the chromatin structure of the adenovirus genome DNA plays critical roles in its genome functions. Previously, we reported that in early phases of infection, incoming viral DNA is associated with both viral core protein VII and cellular histones. Here we show that in late phases of infection, newly synthesized viral DNA is also associated with histones. We also found that the knockdown of CAF-1, a histone chaperone that functions in the replication-coupled deposition of histones, does not affect the level of histone H3 bound on viral chromatin, although CAF-1 is accumulated at viral DNA replication foci together with PCNA. Chromatin immunoprecipitation assays using epitope-tagged histone H3 demonstrated that histone variant H3.3, which is deposited onto the cellular genome in a replication-independent manner, is selectively associated with both incoming and newly synthesized viral DNAs. Microscopic analyses indicated that histones but not USF1, a transcription factor that regulates viral late gene expression, are excluded from viral DNA replication foci and that this is achieved by the oligomerization of the DNA binding protein (DBP). Taken together, these results suggest that histone deposition onto newly synthesized viral DNA is most likely uncoupled with viral DNA replication, and a possible role of DBP oligomerization in this replication-uncoupled histone deposition is discussed.

Section: Cells and Virusessupporting

confidence: 91%

Replication-Uncoupled Histone Deposition during Adenovirus DNA Replication

Komatsu

Nagata

2012

“…Like the herpes simplex virus 1 genome, the HAdV genome was shown to associate with H3.3 (50,51), suggesting that HAdV DNA is chromatinized. The core/DNA complex enters the nucleus and is decondensed prior to the onset of early viral gene transcription (52).…”

Section: Discussionmentioning

confidence: 99%

KAP1 Is a Host Restriction Factor That Promotes Human Adenovirus E1B-55K SUMO Modification

Bürck

Mund

Berscheminski

et al. 2016

Once transported to the replication sites, human adenoviruses (HAdVs) need to ensure decondensation and transcriptional activation of their viral genomes to synthesize viral proteins and initiate steps to reprogram the host cell for viral replication. These early stages during adenoviral infection are poorly characterized but represent a decisive moment in the establishment of a productive infection. Here, we identify a novel host viral restriction factor, KAP1. This heterochromatin-associated transcription factor regulates the dynamic organization of the host chromatin structure via its ability to influence epigenetic marks and chromatin compaction. In response to DNA damage, KAP1 is phosphorylated and functionally inactive, resulting in chromatin relaxation. We discovered that KAP1 posttranslational modification is dramatically altered during HAdV infection to limit the antiviral capacity of this host restriction factor, which represents an essential step required for efficient viral replication. Conversely, we also observed during infection an HAdV-mediated decrease of KAP1 SUMO moieties, known to promote chromatin decondensation events. Based on our findings, we provide evidence that HAdV induces KAP1 deSUMOylation to minimize epigenetic gene silencing and to promote SUMO modification of E1B-55K by a so far unknown mechanism. IMPORTANCEHere we describe a novel cellular restriction factor for human adenovirus (HAdV) that sheds light on very early modulation processes in viral infection. We reported that chromatin formation and cellular SWI/SNF chromatin remodeling play key roles in HAdV transcriptional regulation. We observed that the cellular chromatin-associated factor and epigenetic reader SPOC1 represses HAdV infection and gene expression. Here, we illustrate the role of the SPOC1-interacting factor KAP1 during productive HAdV growth. KAP1 binds to the viral E1B-55K protein, promoting its SUMO modification, therefore illustrating a crucial step for efficient viral replication. Simultaneously, KAP1 posttranslational modification is dramatically altered during infection. We observed an HAdV-mediated decrease in KAP1 SUMOylation, known to promote chromatin decondensation events. These findings indicate that HAdV induces the loss of KAP1 SUMOylation to minimize epigenetic gene silencing and to promote the SUMO modification of E1B-55K by a so far unknown mechanism.

“…Indeed, genome-wide analysis has revealed acetylated histones across the hAd genome (15). These include H3, H3.1, and H3.3, which are acetylated at K9 and K14, presumably by GCN5, and also at K18, which is presumably acetylated by p300/CBP (15,22,41), yet no direct role for E1A or GCN5 in this process was demonstrated in either report. Consistent with previous reports, we show here that E1A is not required for H3 deposition and K9/K14 acetylation, as cells infected with hAd with deletion of E1A (⌬E1A) still acquire both H3 and K9/K14-acetylated H3 (H3-Ac) on the viral E4 promoter (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…GCN5 has a well-established role as a transcriptional coactivator that acetylates histone H3 at K9 and K14 and facilitates transcriptional elongation in the context of the SAGA complex (4,9,32,40). Although the viral genome is initially devoid of cellular histones, recent reports demonstrate that histones, including H3, bind to viral DNA during the early phases of infection (15,22,41). These reports indicated that the H3 associated with the viral genome is acetylated at K9 and K14 during virus infection, likely by GCN5.…”

Section: Figmentioning

confidence: 98%

“…Recent studies have shown that histone H3 associates with the viral genome during infection and that it is acetylated during the course of infection, presumably by GCN5 (15,22,41). Given that the predominant function of GCN5 on cellular chromatin is to acetylate histone H3 on K9 and K14, it is possible that GCN5 is needed to modify cellular histones associated with the hAd genome.…”

Section: E4orf6/7 Gene Expression Is Enhanced In Cells Lacking Gcn5mentioning

confidence: 99%

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Cellular GCN5 Is a Novel Regulator of Human Adenovirus E1A-Conserved Region 3 Transactivation

Ablack

Cohen

Thillainadesan

et al. 2012

The largest isoform of adenovirus early region 1A (E1A) contains a unique region termed conserved region 3 (CR3). This region activates viral gene expression by recruiting cellular transcription machinery to the early viral promoters. Recent studies have suggested that there is an optimal level of E1A-dependent transactivation required by human adenovirus (hAd) during infection and that this may be achieved via functional cross talk between the N termini of E1A and CR3. The N terminus of E1A binds GCN5, a cellular lysine acetyltransferase (KAT). We have identified a second independent interaction of E1A with GCN5 that is mediated by CR3, which requires residues 178 to 188 in hAd5 E1A. GCN5 was recruited to the viral genome during infection in an E1A-dependent manner, and this required both GCN5 interaction sites on E1A. Ectopic expression of GCN5 repressed transactivation by both E1A CR3 and full-length E1A. In contrast, RNA interference (RNAi) depletion of GCN5 or treatment with the KAT inhibitor cyclopentylidene-[4-(4=-chlorophenyl)thiazol-2-yl]hydrazone (CPTH2) resulted in increased E1A CR3 transactivation. Moreover, activation of the adenovirus E4 promoter by E1A was increased during infection of homozygous GCN5 KATdefective (hat/hat) mouse embryonic fibroblasts (MEFs) compared to wild-type control MEFs. Enhanced histone H3 K9/K14 acetylation at the viral E4 promoter required the newly identified binding site for GCN5 within CR3 and correlated with repression and reduced occupancy by phosphorylated RNA polymerase II. Treatment with CPTH2 during infection also reduced virus yield. These data identify GCN5 as a new negative regulator of transactivation by E1A and suggest that its KAT activity is required for optimal virus replication.T he early region 1A (E1A) gene is the first viral gene expressed in cells upon infection with human adenovirus (hAd). Early in infection the primary E1A transcript is alternatively spliced to yield two predominant isoforms that perform two essential functions required to initiate the viral replication cycle. First, E1A uncouples the cell cycle control program of the host cell, driving it into S phase to provide an optimal cellular environment for viral replication. The smaller major E1A isoform (243 residues in human adenovirus type 5 [hAd5] E1A) is sufficient to override cell cycle progression and drive cells into S phase (16,34). The other critical function of E1A is to activate transcription of the early viral promoters. Transactivation of the hAd early genes is predominantly mediated by the product of the largest E1A isoform (6,21,30,31). In hAd5 the 13S E1A mRNA codes for a 289-residue protein that differs from the 243R E1A protein by a unique 46-aminoacid (aa) C4 zinc finger domain located within conserved region 3 (CR3), which is essential for viral transactivation (6, 21). In fact, CR3 alone is sufficient to stimulate transcription of an E2 promoter when microinjected into HeLa cells (25). The region of hAd5 E1A spanning residues 139 to 204 (which includes CR3) is critica...