Dysregulation of Host Cellular microRNA Expression by the Human Papillomavirus E6 and E7 Oncoproteins

Abstract

Human papillomaviruses (HPVs) are small DNA viruses with a tropism for squamous epithelia. Nearly all cervical cancers are caused by a small group of high-risk HPVs and these viruses are also associated with anal, vaginal, vulvar, penile and oropharyngeal cancers. The high-risk HPV16 E6 and E7 proteins are the major drivers of cell transformation and HPV-carcinogenesis. The most well known oncogenic activities of HPV16 E6 and E7 are degradation of the p53 and retinoblastoma tumor suppressors, respectively. However, HPV16 E6/E7 also drive transformation via other mechanisms, and this dissertation investigates how HPV16 E6 and E7 dysregulate expression of non-coding RNAs, with a focus on microRNAs (miRs). Although high-risk HPVs are not known to encode miRs, these viruses have been shown to alter the expression of host cellular miRs. In many previous HPV miR-profiling studies, differentiating cells were analyzed. Given HPVs alter epithelial cell differentiation, it is unclear whether reported changes in miRs are directly caused by HPV gene expression or represent sequelae of HPV-induced changes in epithelial cell differentiation. This dissertation examines cellular miRs modulated by expression of HPV16 E6/E7 in undifferentiated primary human epithelial cells. In Chapter 2, we perform a comprehensive analysis of miRs and mRNAs altered by HPV16 E6/E7 using deep sequencing. Pairing miRs with potential targets, our data show that many observed changes in mRNA expression may be due, in part, to perturbation of miRs by HPV16 E6/E7. In Chapter 3, we investigate the alteration of miR biogenesis enzymes by HPV16 E6/E7. Expression of the HPV16 oncoproteins increases levels of DROSHA and DICER. Furthermore, manipulation of DROSHA levels may be one mechanism by which HPV16 E6/E7 expression dysregulates cellular miR expression. In Chapter 4, we examine how expression of HPV16 E6/E7 alters the expression of miRs in extracellular vesicles. Some miRs are similarly regulated by HPV16 E6/E7 in cells and extracellular vesicles whereas others are not. Overall, this dissertation shows that modulation of miRs is an important part of HPV16 E6/E7 mediated reprogramming of cellular gene expression and may contribute to the transforming activities of these proteins.