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Papillomavirus Genomes Associate with BRD4 to Replicate at Fragile Sites in the Host Genome

2014-05-15 12:12:22

Public Library of Science | Pathogens; 2014 May 15; DOI: 10.1371/journal.ppat.1004117



Moon Kyoo Jang, Kui Shen, Alison A. McBride



Abstract



It has long been recognized that oncogenic viruses often integrate close to common fragile sites. The papillomavirus E2 protein, in complex with BRD4, tethers the viral genome to host chromatin to ensure persistent replication. Here, we map these targets to a number of large regions of the human genome and name them Persistent E2 and BRD4-Broad Localized Enrichments of Chromatin or PEB-BLOCs. PEB-BLOCs frequently contain deletions, have increased rates of asynchronous DNA replication, and are associated with many known common fragile sites. Cell specific fragile sites were mapped in human C-33 cervical cells by FANCD2 ChIP-chip, confirming the association with PEB-BLOCs. HPV-infected cells amplify viral DNA in nuclear replication foci and we show that these form adjacent to PEB-BLOCs. We propose that HPV replication, which hijacks host DNA damage responses, occurs adjacent to highly susceptible fragile sites, greatly increasing the chances of integration here, as is found in HPV-associated cancers.



Author Summary



Papillomavirus cause persistent, but mostly self-limiting, infections of the host epithelium. However, a subset of oncogenic papillomaviruses is the causative agent of certain human cancers. In persistent infection the viral genomes are tethered to host chromosomes to maintain and partition the extrachromosomal viral genomes to daughter cells. However, in cancers viral DNA is often found integrated close to common fragile sites, regions prone to breakage, amplification and deletion. We show that the viral E2 and cellular BRD4 proteins are associated with fragile regions of the human genome and nucleate viral replication foci at these sites. This is a resourceful strategy for a virus that uses the host DNA damage response to amplify viral DNA. However, the outcome may be increased accidental integration of viral DNA, which in the case of the oncogenic viruses can promote carcinogenesis.



Introduction



Papillomaviruses are an ancient group of viruses that establish a persistent infection in the host epithelium. To maintain such a long-term infection, the E2 protein from a subset of papillomaviruses binds to the viral genome and tethers it to the host chromosomes. The bromodomain protein, BRD4, binds to mitotic chromosomes with E2 is essential for regulation of viral transcription and is recruited to early viral replication foci. BRD4 is a mitotic chromosome-associated protein that interacts with acetylated histone tails and is a key regulator of the pTEF-b elongation factor. There has been a recent explosion of data as BRD4 has been implicated in regulation of cell cycle, mitotic memory, transcription of MYC and regulation of viral gene expression. BRD4 is highly enriched at super-enhancers that maintain expression of oncogenes in tumors and is a promising therapeutic target for a number of cancers.



Most HPV infections result in benign lesions, but several are oncogenic and the causative agents of human cancer. Almost all cervical cancer is associated with HPV infection, and oncogenic HPVs are responsible for many anal, penile, vaginal and oropharyngeal cancers. The HPV genome is found integrated into the host genome in over 80% cancers and this promotes malignant progression. The integration event is accidental, but the resulting deregulation of expression of the E6 and E7 oncogenes gives cells a selective growth advantage. There is a predilection for integration within the vicinity of fragile sites.



Papillomaviruses are adept at hijacking host functions and induce a host DNA damage response (DDR) in nuclear foci, resulting in an influx of repair factors that the virus exploits to amplify its own DNA. We show that the HPV E2 protein binds with BRD4 to regions that are highly susceptible to replication stress and overlap many common fragile sites. Common fragile sites are hypersensitive to DNA damage and their replication is often incomplete in the G2 phase of the cell cycle. Thus, they represent a vulnerable and very clever target for papillomavirus replication. Furthermore, replication adjacent to fragile sites may explain the high incidence of integration of oncogenic HPV genomes at these loci.



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