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Published on: November 12, 2015
Disruption of NBS1/MRN Complex Formation by E4orf3 Supports NF-κB That Licenses E1B55K-Deleted Adenovirus-Infected
Nujud Almuzaini1, Madison Moore1, Marjorie Robert-Guroff2
1Department of Biology, College of Arts and Sciences, Howard Universitygrid.257127.4, Washington, D.C., USA.
Abstract:
Cells increase their DNA content greater than the G2/M (DNA > 4n) phases along the path to cancer. The signals that support this increase in DNA content remain poorly understood. Cells infected with adenovirus (Ad) similarly develop DNA > 4n and share a need to bypass the DNA damage response (DDR) signals that trigger cell cycle arrest, and/or cell death. Ads with deletion in early region 1B55K (ΔE1B Ad) are oncolytic agents that are currently being explored for use in vaccine delivery. Interestingly, they promote higher levels of DNA > 4n than Ads that contain E1B55K. Existing in these and almost all Ads that are being explored for clinical use, is early region 4 (E4). The Ad E4 open reading frame 3 (E4orf3) is a viral oncogene that interferes with the ability of cells to respond to DNA damage by disrupting MRN complex formation. Our study reveals that E4orf3 is required for the enhanced fraction of ΔE1B Ad-infected cells with DNA > 4n. For that reason, we explored signaling events mediated by E4orf3. We found that in ΔE1B Ad-infected cells, E4orf3, as reported by others, isolates NBS1 in nuclear dots and tracks. This allows for elevated levels of phosphorylated ATM that is linked to transcriptionally active NF-κB. Pharmacological inhibition of NF-κB reduced the fraction of ΔE1B Ad-infected cells with DNA > 4n while pharmacological inhibition of ATM reduced the levels of nuclear NF-κB and the fraction of ΔE1B Ad-infected cells with DNA > 4n and increased the fraction of dead or dying cells with fragmented DNA. This ability of E4orf3 to disrupt MRN complex formation that allows cells to bypass the cell cycle, evade death, and accumulate DNA > 4n, may be linked to its oncogenic potential. IMPORTANCE Genome instability, a hallmark of cancer, exists as part of a cycle that leads to DNA damage and DNA > 4n that further enhances genome instability. Ad E4orf3 is a viral oncogene. Here, we describe E4orf3 mediated signaling events that support DNA > 4n in ΔE1B Ad-infected cells. These signaling events may be linked to the oncogenic potential of E4orf3 and may provide a basis for how some cells survive with DNA > 4n.
Insights
Adenovirus E4orf3 protein promotes cancer-like DNA increases in infected cells by disrupting DNA damage responses. This viral oncogene
Area of Science:
- Molecular Biology
- Virology
- Oncology
Background:
- Cancer cells exhibit increased DNA content (DNA > 4n), a process poorly understood.
- Adenovirus (Ad) infection can induce DNA > 4n, necessitating bypass of DNA damage response (DDR) pathways.
- Oncolytic Ads with deleted early region 1B55K (ΔE1B Ad) promote higher DNA > 4n levels.
Purpose of the Study:
- To investigate the role of Ad early region 4 open reading frame 3 (E4orf3) in promoting DNA > 4n.
- To elucidate the signaling pathways mediated by E4orf3 in ΔE1B Ad-infected cells.
Main Methods:
- Studied ΔE1B Ad-infected cells, focusing on the viral oncogene E4orf3.
- Investigated E4orf3's effect on MRN complex formation, NBS1 localization, ATM phosphorylation, and NF-κB activity.
- Utilized pharmacological inhibitors of ATM and NF-κB to assess their impact on DNA > 4n and cell death.
Main Results:
- E4orf3 is essential for the increased fraction of ΔE1B Ad-infected cells with DNA > 4n.
- E4orf3 disrupts MRN complex formation, leading to NBS1 nuclear sequestration, elevated phosphorylated ATM, and transcriptionally active NF-κB.
- Inhibition of NF-κB reduced DNA > 4n cells; inhibition of ATM decreased nuclear NF-κB, reduced DNA > 4n cells, and increased cell death.
Conclusions:
- Ad E4orf3 facilitates DNA > 4n accumulation by disrupting the MRN complex, enabling cells to bypass cell cycle arrest and evade death.
- E4orf3-mediated signaling, involving ATM and NF-κB, contributes to genome instability and may underlie its oncogenic potential.
- Understanding these mechanisms offers insight into how cells survive with elevated DNA content, a hallmark of cancer.
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