A multi-platform analysis of the E2-DNA interaction in HPV oncogene regulation

Abstract

The human papillomavirus (HPV) is the most common sexually transmitted infection, with approximately 13 million new infections in the United States each year. While many HPV infections are transient, persistent infections with high-risk strains can lead to various cancers, including cervical, anal, vulvar, and oropharyngeal cancers. HPV is classified into different types, with high-risk strains, such as HPV 16 and 18, being strongly associated with cancer. Despite the development of vaccines that target several high-risk HPV types, there remains no cure for existing infections. HPV's lifecycle and its role in oncogenesis are driven by a complex interplay of viral proteins, including the E2 protein, which is central to viral replication and transcriptional regulation. ☐ The HPV genome consists of a double-stranded DNA of approximately 8 kb, divided into three main regions: the early region, late region, and long control region (LCR). The E2 protein, which is encoded by the early region, plays a crucial role in regulating viral DNA replication and transcription. Specifically, E2 binds to conserved DNA sequences in the LCR, facilitating viral replication and controlling the expression of the viral oncogenes E6 and E7. These oncogenes are responsible for the progression and development of cancer in host cells. E2 also controls transcription by acting as a repressor or activator, depending on its binding affinity and the presence of viral DNA integration. In cases where the E2 gene is disrupted due to viral integration into the host genome, the expression of E6 and E7 becomes deregulated, leading to oncogenic progression. ☐ Despite advances in understanding HPV's molecular mechanisms, gaps remain, particularly regarding the role of the E2-DNA interaction in regulating oncogenesis. This thesis aimed to address three key areas: (i) investigate whether synthetic expression systems can be utilized to study HPV oncogenic regulation, (ii) evaluate the effects of HPV mutations on E2-mediated transcription of the E6/E7 oncogenes across different HPV types, and (iii) assess the utility of various gene expression techniques in enhancing our understanding of the E2-DNA interaction and its role in transcriptional regulation. Through biochemical assays and the study of DNA binding site variants, this research contributes to a deeper understanding of the E2 protein's role in regulating HPV gene expression and oncogenesis. ☐ The findings presented in this thesis offer insights into the dynamic between HPV E2, its DNA binding sites, and the consequences of mutations in these sites. By exploring how the E2-DNA interaction affects transcriptional regulation and oncogene expression in both clinical and experimental contexts, this research provides a foundation for the development of new therapeutic strategies targeting HPV infections and their associated cancers. Understanding these molecular processes is critical for advancing the development of antiviral treatments that could prevent or halt the progression of HPV-induced cancers, addressing a major public health concern globally.