Human Papillomavirus Type 16 E6* Induces Oxidative Stress and DNA Damage

Abstract

High-risk types of human papillomavirus (HPV) are the causative agents of virtually all cases of cervical cancer and a significant proportion of other anogenital cancers, as well as both oral and pharyngeal cancers. The high-risk types encode two viral oncogenes, E6 and E7, which work together to initiate cell transformation. Multiple steps involving the activities and interactions of both viral and cellular proteins are involved in the progression from HPV infection to cell transformation to cancer. The E6 oncoprotein is expressed as several isoforms: a full-length variant referred to as E6 and a few shorter isoforms collectively referred to as E6*. In this study, we found that expression of E6* increased the level of reactive oxygen species (ROS) in both HPVpositive and HPV-negative cells. This increased oxidative stress led to higher levels of DNA damage, as assessed by the comet assay, quantification of 8-oxoguanine, and poly(ADP-ribose) polymerase 1. The observed increase in ROS may be due to a decrease in cellular antioxidant activity, as we found that E6* expression also led to decreased expression of superoxide dismutase isoform 2 and glutathione peroxidase. These studies indicate that E6* may play an important role in virus-induced mutagenesis by increasing oxidative stress and DNA damage. IMPORTANCEOur findings demonstrate for the first time that an HPV gene product, E6*, can increase ROS levels in host cells. This ability may play a significant role both in the viral life cycle and in cancer development, because an increase in oxidative DNA damage may both facilitate HPV genome amplification and increase the probability of HPV16 DNA integration. Integration, in turn, is thought to be an important step in HPV-mediated carcinogenesis.H igh-risk (HR) types of human papillomavirus (HPV) are the causative agents of virtually all cases of cervical cancer as well as a significant percentage of other anogenital and oropharyngeal cancers. In fact, current estimates indicate that HPV infection may be associated with as many as 93% of anal cancers, 63% of oropharyngeal cancers, 40% of penile cancers, 64% of vaginal cancers, and 51% of vulvar cancers (1). HPV infection accounted for approximately 26,700 cases of HPV-related cancers in the United States (2, 3), and it is estimated that 5.2% of all cancers worldwide can be attributed to HPV infection (4). While the incidence of cervical cancer has declined in the last 30 years due to Pap smear screening, the incidence rates of anal, oropharyngeal, and vulvar cancers steadily increased within the same period (1). These numbers underscore the need for ongoing research into the mechanisms behind HPV-related carcinogenesis.The high-risk types of HPV encode two viral oncogenes, E6 and E7, that together serve as the major initiators of cell transformation (5). Multiple steps are involved in the progression from HPV infection to cellular transformation to cancer. Virus-related factors influencing this progression include virus persistence, viral load, and the repr...

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“…This because ROS can promote the DNA oxidation, leading to DNA breaks [5] observed in papillomavirus-infected cells lines (table 1, figure 7), as well as those described in literature [10][11][12][13][14]19]. Similar results were also verified in HPV infected cells, in which it was demonstrated that the HPV-16 E6*-induced oxidative stress and DNA breaks, which are necessary to HPV integration [15,16]. Based on these data, we analyzed the action of BPV-1 E6 recombinant oncoprotein in glycolytic metabolism.…”

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“…However, BPV-infected cells do not reduce the ROS production, even after malignant transformation. Although controversial, these data can be attributed to E6 oncoprotein, which reduces SOD2 expression levels, increasing ROS formation [145]. This action increase the pro-oxidant stimulus (ROS production) and reduces the anti-oxidant cell capability (SOD2), leading to oxidant stress.…”

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“…[85] Additionally, ROS activates HPV [16] and its E6 protein can in turn cause production of more ROS and DNA damage. [86] The E2 protein of HPV also results in production of ROS by interacting with the cells' mitochondria [87] creating oxidized cytosines that can spontaneously deaminate and become thymines. Cytosine to thymine transition mutations can also occur via viral activation of the APOBEC system resulting in epigenetic methylation and deamination of cytosines.…”