Human papillomavirus induces epigenetic reprogramming of human keratinocytes

Diyaf, Afaf Ayad Hadi (2021). Human papillomavirus induces epigenetic reprogramming of human keratinocytes. University of Birmingham. Ph.D.

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Abstract

In this thesis, I investigate the contribution of human papillomavirus (HPV) induced epigenetic changes to the process of cervical carcinogenesis. I used an in vitro progression model of cervical cancer involving primary human foreskin keratinocytes in conjunction with human cervical intraepithelial neoplasia cell line and different cervical cancer cell lines. The Primary Human Foreskin Keratinocyte from one donor transfected with episomal HPV16 and HPV18. Following propagation of PHFK HPV18 and PHFK HPV 16 cell lines, I have shown HPV-induced cytopathic changes in human keratinocytes using immunocytochemistry. I found that HPV-associated with changes in the expression of key epigenetic regulators that follow the HPV life cycle and the process of cervical carcinogenesis. Using PCR methods I showed that specific regions of HPV16 and HPV18 genome are prone to disruption during HPV integration. Overexpression of HPV E7 oncoprotein precedes the disruption of full length HPV E2 and induces expression and enzymatic activity of DNMT1 and DNMT3B. Using co- immunoprecipitation, I found that the endogenous HPV E7 oncoprotein interacts at physiological level with DNA methyltransferase 3B. I have also shown, using chromatin immunoprecipitation, that HPV associated with pattern of epigenetic switching from histone 3 lysine 27 trimethylation to DNA methylation. Using a genome wide methylation array, I found that HPV18 integration induces changes to the human DNA methylome similar to the changes expected in cervical neoplasia. Finally, I show that methylated tumour suppressor genes following HPV integration are reversibly demethylated with 5-azacitidine.

Type of Work:

Thesis (Doctorates > Ph.D.)

Award Type:

Doctorates > Ph.D.

Supervisor(s):