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Adenovirus proteins regulate nuclear sumoylation and hijack the chromatin remodeling complex PRC1 to mediate silencing of anti-viral genes

  • Author(s): Higginbotham, Jennifer
  • Advisor(s): O'Shea, Clodagh
  • Desai, Arshad
  • et al.
Abstract

Adenovirus E4-ORF3 and E1B-55K converge in subverting critical overlapping cellular pathways to facilitate virus replication. Here, we show that E1B-55K and E4-ORF3 induce sumoylation and the assembly of SUMO2/3 viral genome replication domains. Using a conjugation-deficient SUMO2 construct, we demonstrate that SUMO2/3 is recruited to E2A viral genome replication domains through noncovalent interactions. E1B-55K and E4-ORF3 have critical functions in inactivating MRN and ATM to facilitate viral genome replication. We show that ATM kinase inhibitors rescue ΔE1B-55K/ΔE4-ORF3 viral genome replication and that the assembly of E2A domains recruits SUMO2/3 independently of E1B-55K and E4-ORF3. However, the morphology and organization of SUMO2/3-associated E2A domains is strikingly different from that in wild-type Ad5-infected cells. These data reveal that E1B-55K and E4-ORF3 specify the nuclear compartmentalization and structure of SUMO2/3-associated E2A domains, which could have important functions in viral replication. We show that E4-ORF3 specifically targets and sequesters the cellular E3 SUMO ligase PIAS3 and the SUMO proteases SENP1 and SENP2. The assembly of E4-ORF3 into a multivalent nuclear matrix is required to target PIAS3, SENP1, and SENP2. In contrast to MRN, PIAS3, SENP1, and SENP2 are targeted by E4-ORF3 proteins from disparate adenovirus subgroups. Our studies reveal that PIAS3, SENP1, and SENP2 are novel and evolutionarily conserved targets of E4-ORF3 in human adenovirus infections. Furthermore, we reveal that viral proteins not only disrupt but also usurp SUMO2/3 to transform the nucleus and assemble novel genomic domains that could facilitate pathological viral replication. We also show that adenovirus protein E2A, which is paramount for viral genome replication, is itself sumoylated and has SUMO-interacting motifs, which may play a role in adenovirus replication.

The Polycomb Repressive Complex 1 (PRC1) has been shown to be required to maintain the transcriptionally repressive state of many genes, acting via both chromatin remodeling and modification of histones. We have identified PRC1 complex members CBX2, CBX4, and Ring1b as novel cellular targets mislocalized with E4-ORF3 into the nuclear scaffold that specifies heterochromatin silencing of p53 and antiviral target genes. These studies help to understand how transcription of cellular genes is misregulated in adenovirus infected cells and which viral proteins are responsible.

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