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The Human Papillomavirus Type 16 E6 Oncogene Regulates microRNA-218 via the Histone Acetyltransferase p300 in Cervical Carcinoma Cells

Gardiner, Amy Sabrina (2010) The Human Papillomavirus Type 16 E6 Oncogene Regulates microRNA-218 via the Histone Acetyltransferase p300 in Cervical Carcinoma Cells. Doctoral Dissertation, University of Pittsburgh. (Unpublished)

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Abstract

Human papillomaviruses (HPVs) are involved in the pathogenesis of different types of human cancers, especially cancer of the cervix (CaCx). MicroRNAs (miRNAs) are post-transcriptional gene regulators that have recently been associated with many types of cancers. We analyzed the expression of cellular miRNAs in HPV-16 positive cervical cell lines and tissues via microarray, Northern blotting, and quantitative RT-PCR. Three miRNAs were overexpressed and 24 underexpressed in cervical cell lines containing integrated HPV-16 DNA compared to the normal cervix. An HPV-negative CaCx cell line, C-33A, showed underexpression of four miRNAs compared to the normal cervix. Also, nine miRNAs were overexpressed and only one underexpressed in cell lines containing integrated HPV-16 DNA compared to C-33A. MicroRNA-218 (miR-218) was specifically underexpressed in cell lines, cervical lesions and cancer tissues containing integrated HPV-16 DNA compared to both C-33A and the normal cervix. Exogenous expression of the HPV-16 E6 oncogene reduced miR-218 expression, and conversely, RNA interference of E6/E7 oncogenes in an HPV-16 positive cell line increased miR-218 expression. Furthermore, exogenous expression of miR-218 in HPV-16 positive cell lines decreased expression of the epithelial-specific gene LAMB3, which is involved in cell migration and tumorigenicity. We analyzed the expression of cellular miRNAs in additional clinical samples including six HPV-16 positive cervical cancers, three cervical dysplasias, and four normal cervical tissues using TaqMan® MicroRNA Arrays V2.0. Eighteen miRNAs were overexpressed and two underexpressed in cervical cancer tissues compared to normal cervical tissues. Nine miRNAs were consistently overexpressed, and two miRNAs were consistently underexpressed, including miR-218, in cervical dysplasias and cervical cancer tissues compared to normal tissues. We also found that exogenous expression of miR-218 in HPV-16 positive cell lines decreased expression of the extracellular matrix protein MMP3, which is involved in the epithelial-mesenchymal transition. We also demonstrated that HPV-16 E6 regulates miR-218 via the histone acetyltransferase p300 in cervical cells. The expression of miR-218 was found to be reduced in HPV-negative cells upon p300 knock-down or p300 inhibition with anacardic acid, and expression of miR-218 was reactivated in HPV-16 positive cells upon EZH2 knock-down or inhibition with adenosine dialdehyde. Reactivation of miR-218 was enhanced upon treatment with a combination of 5-azacytidine, trichostatin A, and adenosine dialdehyde. We also demonstrated that miR-218 reduces the migration and invasion of SiHa cervical cancer cells, indicating that miR-218 functions as a tumor suppressor in cervical cancer and that it may have therapeutic value.


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Details

Item Type: University of Pittsburgh ETD
Status: Unpublished
Creators/Authors:
CreatorsEmailPitt UsernameORCID
Gardiner, Amy Sabrinaasg18@pitt.eduASG18
ETD Committee:
TitleMemberEmail AddressPitt UsernameORCID
Committee ChairKhan, Saleemkhan@pitt.eduKHAN
Committee MemberJohn, Binobino@pitt.eduBINO
Committee MemberPipas, Jamespipas@pitt.eduPIPAS
Committee MemberSteinman, Richardsteinman@pitt.eduSTEINMAN
Committee MemberDuensing, Stefanduensing@pitt.eduDUENSING
Date: 23 December 2010
Date Type: Completion
Defense Date: 10 December 2010
Approval Date: 23 December 2010
Submission Date: 22 December 2010
Access Restriction: No restriction; Release the ETD for access worldwide immediately.
Institution: University of Pittsburgh
Schools and Programs: School of Medicine > Molecular Virology and Microbiology
Degree: PhD - Doctor of Philosophy
Thesis Type: Doctoral Dissertation
Refereed: Yes
Uncontrolled Keywords: cervical cancer; human papillomavirus; microRNA
Other ID: http://etd.library.pitt.edu/ETD/available/etd-12222010-162351/, etd-12222010-162351
Date Deposited: 10 Nov 2011 20:11
Last Modified: 15 Nov 2016 13:55
URI: http://d-scholarship.pitt.edu/id/eprint/10452

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