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The DNA repair protein, O6-Methylguanine-DNA methyltransferase is a proteolytic target for the E6 human papillomavirus oncoprotein

Oncogene volume 21, pages 5940–5945 (2002)Cite this article

Abstract

We have previously shown that O6-methylguanine-DNA methyltransferase (MGMT), a DNA repair protein that protects tissues against toxic and carcinogenic effects of alkylating agents, is degraded through ubiquitination-dependent proteolysis. Here, we investigated the role of the human papillomavirus (HPV) E6 protein in MGMT degradation. In three pairs of isogenic human tumor cell lines in which a member of each pair expressed the E6 protein through stable transfection (HCT116/HCT116-E6, MCF7/MCF7-E6, and RKO/RKO-E6), we found a consistent 40–55% reduction in the MGMT protein level and its activity in all E6-expressing cells compared with the parent cells (P=<0.05). E6 expression did not, however, alter the levels of MGMT mRNA. Addition of the recombinant MGMT (rMGMT) protein to extracts of HCT116/E6 cells resulted in the binding of E6 to MGMT. Further, the purified E6 protein promoted the degradation of rMGMT in rabbit reticulocyte lysates. Immunoprecipitation assays showed the presence of a ternary protein complex between MGMT, E6, and the cellular ubiquitin-ligase E6-associated protein (E6-AP). Transient transfection of the p53-null H1299 lung tumor cells with an E6 construct also down-regulated the MGMT. The MGMT protein also showed structural features that are compatible for interaction with the E6, and E6-AP components. Collectively, these data suggest that the oncogenic E6 proteins enhance the ubiquitin-dependent proteolysis of MGMT.

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Abbreviations

MGMT:

O6-methylguanine-DNA methyltransferase

HPV:

human papillomavirus

BCNU:

1,3-bis(2-chloroethyl)-1-nitrosourea

rMGMT:

recombinant MGMT protein

E6-AP:

E6-Associated Protein

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Acknowledgements

We thank Dr Jiang Shou for initial experiments and technical assistance. We are grateful to Drs Michael Kastan, Albert Fornace Jr, and Wafik el-Deiry for supply of E6-transfected cell lines. This work was supported by grants from the National Institutes of Health (CA-74321), Pediatric Brain Tumor Foundation of the United States, and the Association for Reseach of Childhood Cancer to KS Srivenugopal.

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  1. Department of Neurosurgery, Section of Molecular Therapeutics, The University of Texas MD Anderson Cancer Center, Houston, 77030-4009, Texas, TX, USA

    Kalkunte S Srivenugopal & Francis Ali-Osman

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  2. Francis Ali-Osman
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Correspondence to Kalkunte S Srivenugopal.

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Srivenugopal, K., Ali-Osman, F. The DNA repair protein, O6-Methylguanine-DNA methyltransferase is a proteolytic target for the E6 human papillomavirus oncoprotein. Oncogene 21, 5940–5945 (2002). https://doi.org/10.1038/sj.onc.1205762

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