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Use of Interferon-γ Enzyme-linked Immunospot Assay to Characterize Novel T-cell Epitopes of Human Papillomavirus
Published on: March 8, 2012
HPV 16 E6 blocks TNF-mediated apoptosis in mouse fibroblast LM cells
P J Duerksen-Hughes1, J Yang, S B Schwartz
1Department of Biology, Georgia State University, Atlanta, Georgia 30302, USA. biopjd@panther.gsu.edu
Abstract:
The interaction between hosts and the viruses that infect them is a dynamic one, and a growing literature documents the fact that many viruses have developed mechanisms designed to avoid elimination by the host immune system. One of the immune strategies used by the host and targeted by virus proteins is apoptosis triggered by the cytokine tumor necrosis factor (TNF). Mouse fibroblast LM cells are spontaneously sensitive to TNF. When the wild-type E6 protein from the human papillomavirus type 16 (HPV 16) was expressed in LM cells, the cells became resistant to TNF. This resistance was examined by several means, including cell morphology, the dose- and time-independent response to TNF in a cell death ELISA, trypan blue exclusion, and cell proliferation. The level of p53 did not rise in TNF-treated cells prior to apoptosis, suggesting a p53-independent mechanism. Significant, though not complete, resistance to TNF was also observed following transfection of a plasmid expressing a mutant E6 protein, which is unable to mediate rapid degradation of the p53 tumor suppressor. These results indicate that the HPV 16 E6 protein can protect LM cells from TNF-triggered apoptosis and likely does so by a mechanism other than mediation of p53 degradation.
Insights
Human papillomavirus type 16 E6 protein confers resistance to tumor necrosis factor (TNF)-induced apoptosis in mouse fibroblast cells. This protection occurs independently of the p53 tumor suppressor, suggesting a novel viral immune evasion strategy.
Area of Science:
- Virology
- Immunology
- Cell Biology
Background:
- Host-virus interactions involve viral mechanisms to evade host immune responses.
- Cytokine tumor necrosis factor (TNF)-induced apoptosis is a key host immune strategy targeted by viruses.
- Mouse fibroblast LM cells are sensitive to TNF-induced cell death.
Purpose of the Study:
- To investigate the effect of human papillomavirus type 16 (HPV 16) E6 protein on TNF-induced apoptosis in LM cells.
- To determine the mechanism by which HPV 16 E6 protein confers resistance to TNF.
Main Methods:
- Expression of wild-type and mutant HPV 16 E6 protein in LM cells.
- Assessment of TNF-induced apoptosis using cell morphology, cell death ELISA, trypan blue exclusion, and cell proliferation assays.
- Analysis of p53 levels in TNF-treated cells.
Main Results:
- Expression of HPV 16 E6 protein rendered LM cells resistant to TNF-induced apoptosis.
- Resistance to TNF was observed even with a mutant E6 protein unable to degrade p53.
- p53 levels did not increase in TNF-treated cells prior to apoptosis, indicating a p53-independent mechanism.
Conclusions:
- The HPV 16 E6 protein protects LM cells from TNF-triggered apoptosis.
- This protection is likely mediated by a mechanism independent of p53 degradation.
