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PLoS One. 2013 May 30;8(5):e64781. doi: 10.1371/journal.pone.0064781. Print 2013.

An RNA aptamer provides a novel approach for the induction of apoptosis by targeting the HPV16 E7 oncoprotein.

Author information

1
School of Molecular and Cellular Biology, Faculty of Biological Sciences and Astbury Centre for Structural Molecular Biology, University of Leeds, Leeds, United Kingdom.

Abstract

BACKGROUND:

Human papillomavirus 16 (HPV16) is a high-risk DNA tumour virus, which is a major causative agent of cervical cancer. Cellular transformation is associated with deregulated expression of the E6 and E7 oncogenes. E7 has been shown to bind a number of cellular proteins, including the cell cycle control protein pRb. In this study, RNA aptamers (small, single-stranded oligonucleotides selected for high-affinity binding) to HPV16 E7 were employed as molecular tools to further investigate these protein-protein interactions.

METHODOLOGY/PRINCIPAL FINDINGS:

This study is focused on one aptamer (termed A2). Transfection of this molecule into HPV16-transformed cells resulted in inhibition of cell proliferation (shown using real-time cell electronic sensing and MTT assays) due to the induction of apoptosis (as demonstrated by Annexin V/propidium iodide staining). GST-pull down and bead binding assays were used to demonstrate that the binding of A2 required N-terminal residues of E7 known to be involved in interaction with the cell cycle control protein, pRb. Using a similar approach, A2 was shown to disrupt the interaction between E7 and pRb in vitro. Furthermore, transfection of HPV16-transformed cells with A2 appeared to result in the loss of E7 and rise in pRb levels, as observed by immunoblotting.

CONCLUSIONS/SIGNIFICANCE:

This paper includes the first characterisation of the effects of an E7 RNA aptamer in a cell line derived from a cervical carcinoma. Transfection of cells with A2 was correlated with the loss of E7 and the induction of apoptosis. Aptamers specific for a number of cellular and viral proteins have been documented previously; one aptamer (Macugen) is approved for clinical use and several others are in clinical trials. In addition to its role as a molecular tool, A2 could have further applications in the future.

PMID:
23738000
PMCID:
PMC3667794
DOI:
10.1371/journal.pone.0064781
[Indexed for MEDLINE]
Free PMC Article

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