HPV16 E2 Is an Immediate Early Marker of Viral Infection, Preceding E7 Expression in Precursor Structures of Cervical Carcinoma

Abstract

The viral E2 gene product plays a crucial role in the human papillomavirus (HPV) vegetative cycle by regulating both transcription and replication of the viral genome. E2 is a transcriptional repressor of the E6 and E7 viral oncogenes for HPV types 16 and 18, which are involved in cervical cancers. Using new polyclonal antibodies against the HPV16 E2 protein, we showed that E2 is expressed at various precursor stages of cervical carcinoma by immunohistochemistry on paraffin-embedded clinical samples. E2 was found to be highly expressed in the nuclei and cytoplasm of cells forming the intermediate and upper layers of cervical intraepithelial neoplasia (CIN). We could show that the expressions of E2 and p16INK4a (surrogate marker for oncogenic E7 expression) were exclusive in most of the cases, thus implying that E2 is not expressed together with high levels of E7. Moreover, we found that E2 is expressed in a subset of columnar cells adjacent to the CIN. We could show that expression of E2 is topologically distinct from the proliferation markers p63 and Ki67, whereas it coincides with the expression of cytokeratin K13, a marker of squamous cell differentiation. Expression of E2 also topologically coincides with episomal amplification of viral genomes in the upper layers of CIN1. These in vivo data thus validate previous assumptions of the crucial role of E2 in the early steps of HPV infection and of its negative link with expression of the viral E6 and E7 oncogenes. Cancer Res; 70(13); 5316-25. ©2010 AACR.

Human Papillomavirus E2 Regulates SRSF3 (SRp20) To Promote Capsid Protein Expression in Infected Differentiated Keratinocytes

“…The data support the hypothesis that HR-HPV E2, which is expressed at highest levels in differentiating HPV-infected epithelial cells (5,6), controls expression of SR pro- teins. To test this directly, we compared levels of SR proteins in normal foreskin keratinocytes (NFKs) stably transfected with wild-type HPV genomes or with genomes containing an inactivating point mutation in the transactivation domain of E2.…”

“…Differentiation of infected epithelial cells leads to activation of early and then late gene expression (4). The viral replication and transcription factor, E2, is expressed at greatest levels in the middle to upper layers of the epithelium (5,6) where, together with the viral DNA helicase, E1, it facilitates vegetative viral DNA replication, leading to production of thousands of viral genome copies (7). The viral late proteins, including the capsid proteins L1 and L2, are synthesized in the uppermost granular layer of the epithelium to encapsidate the newly replicated genomes (4).…”

Human Papillomaviruses Oncoproteins

“…E2HPV is involved in viral transcription and replication [116,117], forming together with E1 a complex with viral origin of replication and recruiting cellular DNA replication machinery (DNA polymerases, replication protein A, replication protein C, topoisomerase I/II and proliferating-cell nuclear antigen) in order to facilitate viral DNA replication [116]. It was noted that E2 protein is expressed at relatively high levels in differentiated cells of the intermediate layers of CIN lesions; on the other hand, its expression is decreased with progression of the lesions and is absent in most of the cancers in situ, being inversely correlated with expression of E7 [13,118]. E2 is an unstable protein expressed in both the nuclei and cytoplasm of infected cells, and is degraded through the proteasome [119,120].…”

Oligonucleotide Applications for the Therapy and Diagnosis of Human Papillomavirus Infection

“…The relationship between cervical cancer and high-risk HPVs is related to the integration of viral DNA to the host genome with the disruption of the viral regulator E2 (Pett & Coleman, 2007;Vernon et al, 1997), and to the production of viral proteins E6 and E7 which are sufficient and necessary to acquire and maintain a transformed phenotype (Pirisi et al, 1988;Xue et al, 2010). Because E2 is usually absent in cervical tumor cells and E6/E7 genes are retained and expressed in most tumors, these features are often referred to as the hallmarks of cervical cancer .…”

“…During its life cycle, HPV expresses proteins according the cellular differentiation program that is modified after E2 disruption and the onset of malignant transformation (Pett & Coleman, 2007;Xue et al, 2010). Such features offer several protein targets for the detection of HPV infection and molecular diagnosis of cervical cancer by using aptamers as diagnostic oligonucleotides.…”