Functional Study of HPV16 Minor Capsid Protein L2 in Global Cellular Regulation

Human papillomavirus type 16 minor capsid protein L2 has been shown to assist in the initial entry and intracellular trafficking events leading to nuclear translocation of the viral genome. During my investigations of L2 function, I observed that expression of L2 in a keratinocyte cell line (HaCaT) resulted in phenotypic changes of the cell. In this dissertation, I present data that expression of the L2 protein in this cellular model system HaCaTs resulted in a shift from G0/G1 phase to mitotic S phase, as well as a reduced amount of retinoblastoma protein (Rb) and an increase in Cdc2 phosphorylation. I performed genome-wide host cell mRNA sequencing and identified 2586 differentially expressed genes upon HPV16 L2 expression. Via machine learning and protein network analysis, genes involved in cellular differentiation and proliferation were highlighted as impacted by L2. I also performed the chromatin immunoprecipitation with cellular transcription factor TBX3 and HA-tag-fused L2 in both L2 expressing and absent HaCaT cells. The results demonstrated a reduced binding of TBX3 on the promoter region of its target gene, CDKN1A, when L2 was present. However, this effect of L2 was not observed with another target gene, CDH1, of TBX3, suggesting a complex regulatory effect of L2 on the transcription repression function of TBX3. My results also indicated that other cellular proteins and transcription factors might mediate the alteration of L2 on the TBX3 binding. My findings have implications for the role of L2 at the viral production stages when the virus needs to prevent cellular differentiation while maintaining the cells’ ability to replicate DNA, and might partially compensate the function of HPV16 E7 protein, when E7 is no longer expressed in the infected cells, on maintaining cell dividing and DNA replication. My study suggests a potential novel function of the L2 protein, as a regulator of cellular gene transcription. These insights provided by my study will help to answer the question that how and how much does L2 regulate HPV 16 viral production, as well as suggest means of studying the biology and reproduction of other HPV types and BPVs. This is the first time this question is asked with attempts of assessing. Further understanding of the process of viral production will significantly help with the development of therapeutic methods to clear established infection and to prevent re-infection of the same or different tissues of patients