Escape of p53 protein from E6-mediated degradation in HeLa cells after cisplatin therapy.
We previously reported that therapy of human cervical carcinoma HeLa cells with CP induced segregation of nucleoli and changes of nuclei characteristic of apoptosis. We raised the question of whether p53 can be reactivated by chemotherapy in HeLa cells despite the presence of HPV-encoded E6 activity. Cellular levels of p53 protein increased after CP treatment, reaching a maximum after 6 hr. p53 protein accumulated preferentially in the nucleoli, with a peak after 15 hr. CP-induced nucleolar targeting of p53 appears to be selective because p73, another member of the p53 gene family, accumulated primarily in nuclei in response to CP. Monitoring of the intranuclear distribution of Hdm-2, a negative regulator of p53, revealed this protein in the nucleoli of untreated controls translocated into chromatin during CP therapy. Interestingly, p14(ARF) showed an inverse intranuclear redistribution. Proteasome inhibitors were not able to mimic the effect of CP on p53 levels. Since the reduced stability of wild-type p53 protein in HeLa cells is a consequence of its enhanced ubiquitination by virally encoded E6 protein, resulting in its accelerated degradation, we checked the cellular level of E6 during CP therapy. Six hours after application of CP, E6 protein expression was markedly reduced. This coincided with the increase of cellular p53 and preceded the nucleolar accumulation of p53 protein, indicating that repression of virally coded E6 protein by CP contributes to the restoration of p53 expression.
Wesierska-Gadek
Józefa
J
Institute of Cancer Research, Faculty of Medicine, University of Vienna, Vienna, Austria. Jozefa.Antonia.Gadek-Wesierski@univie.ac.at
Schloffer
Daniela
D
Kotala
Vladimir
V
Horky
Marcel
M
D017209:Apoptosis / Q000187:drug effects*; D002466:Cell Nucleolus / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D002945:Cisplatin / Q000494:pharmacology*; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D004268:DNA-Binding Proteins; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D006367:HeLa Cells; D006801:Humans; D009363:Neoplasm Proteins / Q000378:metabolism; D009687:Nuclear Proteins; D009856:Oncogene Proteins, Viral / Q000037:antagonists & inhibitors / Q000378:metabolism*; D021381:Protein Transport / Q000187:drug effects; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051736:Proto-Oncogene Proteins c-mdm2; D025803:Tumor Suppressor Protein p14ARF / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D015854:Up-Regulation / Q000187:drug effects
D004268:DNA-Binding Proteins; C052603:E6 protein, Human papillomavirus type 18; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D009856:Oncogene Proteins, Viral; D011518:Proto-Oncogene Proteins; D025803:Tumor Suppressor Protein p14ARF; D016159:Tumor Suppressor Protein p53; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2; D002945:Cisplatin