The Adenoviral Oncogene E1A-13S Interacts with a Specific Isoform of the Tumor Suppressor PML To Enhance Viral Transcription

Abstract

PML nuclear bodies (PML NBs), also called ND10, are matrix-bound nuclear structures that have been implicated in a variety of functions, including DNA repair, transcriptional regulation, protein degradation, and tumor suppression. These domains are also known for their potential to mediate an intracellular defense mechanism against many virus types. This is likely why they are targeted and subsequently manipulated by numerous viral proteins. Paradoxically, the genomes of various DNA viruses become associated with PML NBs, and initial sites of viral transcription/replication centers are often juxtaposed to these domains. The question is why viruses start their transcription and replication next to their supposed antagonists. Here, we report that PML NBs are targeted by the adenoviral (Ad) transactivator protein E1A-13S. Alternatively spliced E1A isoforms (E1A-12S and E1A-13S) are the first proteins expressed upon Ad infection. E1A-13S is essential for activating viral transcription in the early phase of infection. Coimmunoprecipitation assays showed that E1A-13S preferentially interacts with only one (PML-II) of at least six nuclear human PML isoforms. Deletion mapping located the interaction site within E1A conserved region 3 (CR3), which was previously described as the transcription factor binding region of E1A-13S. Indeed, cooperation with PML-II enhanced E1A-mediated transcriptional activation, while deleting the SUMO-interacting motif (SIM) of PML proved even more effective. Our results suggest that in contrast to PML NB-associated antiviral defense, PML-II may help transactivate viral gene expression and therefore play a novel role in activating Ad transcription during the early viral life cycle. Human adenovirus type 5 (Ad5) early region 1A (E1A) is the first protein expressed upon infection and plays an essential role in transcriptional activation and induction of cell cycle progression (1). At early times of infection, two major E1A proteins, E1A-12S (243R) and E1A-13S (289R), are synthesized from alternatively spliced mRNA transcripts of the E1A gene (2, 3). The proteins are identical except for a 46-amino-acid (aa) conserved region (CR3) that is unique to the larger E1A protein (3). Despite being very similar, these proteins show significant differences in their biological activities. The larger E1A isoform (E1A-13S) is considered to be primarily responsible for transactivating viral gene expression by interacting with a wide range of transcription factors via its CR3 (4-8).The PML (promyelocytic leukemia) protein was first described in acute promyelocytic leukemia (APL), showing that PML was fused to retinoic acid receptor alpha (RAR␣) due to a chromosomal translocation (t15;17) (9-20). The PML gene encodes multiple isoforms, derived from alternative mRNA splicing, differing only in the C-terminal parts of the proteins (21). PML seems to be responsible for the assembly of nuclear bodies by recruiting other constitutive components such as Daxx (death domain-associated protein), Sp100 (speckled protein ...

Promyelocytic leukemia protein isoform II inhibits infection by human adenovirus type 5 through effects on HSP70 and the interferon response

“…Taken together, these observations support the idea that the E4 Orf3 interaction with PML-II opposes antiviral responses so as to favour productive viral infection, which fits well with the more recent finding that PML-II is needed for a robust type 1 IFN response (Chen et al, 2015). However, it has also been reported that PML-II serves a positive function during Ad5 infection (Berscheminski et al, 2013).…”

“…Previously, Berscheminski and colleagues reported that PML-II was beneficial to Ad5 infection, in apparent contradiction to our findings (Berscheminski et al, 2013). They showed that PML-II potentiated transcriptional activation of a viral early promoter by the E1A 13S protein and that, in a cellular context where all PML proteins were depleted, the addition of exogenous PML-II enhanced virus yield about 3-fold.…”

“…In a PML-null cell, functions observed for added PML-II will be essentially those of its soluble nuclear form. Indeed, a mutated form of PML-II with reduced ability to associate with PML-NBs was more active than the wildtype in cooperating with E1A in the presence of endogenous PML (Berscheminski et al, 2013), suggesting that interactions with other PML isoforms limit this activity. The overexpressed PML-II will also potentially exceed the capacity of E4 Orf3 expressed upon infection to bind and inactivate it.…”

Adenovirus E1A Targets the DREF Nuclear Factor To Regulate Virus Gene Expression, DNA Replication, and Growth

“…It is possible that this interaction plays some role in E1A-mediated transcriptional regulation. An interaction between PML-II and CR3 of E1A was shown to be important for the ability of E1A to activate viral promoters (44), and since DREF is colocalized to PML bodies the two may be linked. Significantly, we observed reduced viral gene expression upon knockdown of DREF (Fig.…”

The SP100 component of ND10 enhances accumulation of PML and suppresses replication and the assembly of HSV replication compartments

“…A central and as-yet unresolved question is whether PML or SP100 plays a role in the early transcription of viral genes after infection (14,42,(47)(48)(49)(50)(51). Both PML and SP100 are degraded within the first 6 hpi, and thus the major effect of PML or SP100 on the transcription of viral DNA would occur during that period (11).…”