Functional Consequences of Directed Mutations in Human Papillomavirus E6 Proteins: Abrogation of p53-Mediated Cell Cycle Arrest Correlates with p53 Binding and Degradation in Vitro

Slebos, Robbert J.C.; Kessis, Theodore D.; Chen, Alex W.; Han, Sung Min; Hedrick, Lora; Cho, Kathleen R.

doi:10.1006/viro.1995.1134

Cited by 21 publications

(18 citation statements)

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“…Furthermore, our results show that HPV5, -6, -8, -11, -42, and -83 do not promote p53 degradation. Although most of these genotypes were previously found to be innocuous to p53 protein expression (11,20,52,69), our report is the first to show the inability of HPV83 E6 to degrade p53. HPV83 was originally isolated from a mouse xenograft system, and its association with cancer remains unclear (7).…”

Section: Discussioncontrasting

confidence: 46%

“…HPV16 E6 was shown to interact in vitro with the karyopherins ␣2, ␤1, and ␤2 (37), suggesting that E6 is actively imported in the nucleus. Our subcellular localization of the 29 HPV E6 proteins used in this study, as fusions with YFP, showed that the E6 proteins from HPV types 11,16,18,26,30,31,33,34,35,39,42,45,51,52,53,56,58,59,66,68,69,70,73,82,83, and 97 accumulate predominantly in the nucleus after transfection (Fig. 5).…”

Section: Discussionmentioning

confidence: 67%

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p53 Degradation Activity, Expression, and Subcellular Localization of E6 Proteins from 29 Human Papillomavirus Genotypes

Mésplède

Gagnon

Bergeron-Labrecque

et al. 2012

J Virol

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Human papillomaviruses (HPVs) are the etiological agents of cervical cancer and other human malignancies. HPVs are classified into high-and low-risk genotypes according to their association with cancer. Host cell transformation by high-risk HPVs relies in part on the ability of the viral E6 protein to induce the degradation of p53. We report the development of a cellular assay that accurately quantifies the p53 degradation activity of E6 in vivo, based on the fusion of p53 to Renilla luciferase (RLuc-p53). This assay was used to measure the p53 degradation activities of E6 proteins from 29 prevalent HPV types and variants of HPV type 16 (HPV16) and HPV33 by determining the amount of E6 expression vector required to reduce by half the levels of RLuc-p53 (50% effective concentration [EC 50 ]). These studies revealed an unexpected variability in the p53 degradation activities of different E6 proteins, even among active types whose EC 50 s span more than 2 log units. Differences in activity were greater between types than between variants and did not correlate with differences in the intracellular localization of E6, with most being predominantly nuclear. Protein and mRNA expression of the 29 E6 proteins was also examined. For 16 high-risk types, spliced transcripts that encode shorter E6*I proteins of variable sizes and abundances were detected. Mutation of the splice donor site in five different E6 proteins increased their p53 degradation activity, suggesting that mRNA splicing can limit the activity of some highrisk E6 types. The quantification of p53 degradation in vivo represents a novel tool to systematically compare the oncogenic potentials of E6 proteins from different HPV types and variants.

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Section: Discussioncontrasting

confidence: 46%

Section: Discussionmentioning

confidence: 67%

p53 Degradation Activity, Expression, and Subcellular Localization of E6 Proteins from 29 Human Papillomavirus Genotypes

Mésplède

Gagnon

Bergeron-Labrecque

et al. 2012

J Virol

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“…In contrast, because of the critical roles of E6 and E7 proteins in the early stages of infection, ribozymes feasibly may be used as antiviral agents in low grade, HPV-16-positive, precancerous lesions. Therefore, ribozymes that target E6 (i.e., R434) are effective mainly in preventing the interaction of E6 with p53 which is reported to be responsible for chromosomal instability and a high mutation rate in infected cells (55)(56)(57).…”

Section: Discussionmentioning

confidence: 99%

Inhibition of HPV-16 E6/E7 immortalization of normal keratinocytes by hairpin ribozymes

Álvarez-Salas

Cullinan²,

Siwkowski

et al. 1998

Proc. Natl. Acad. Sci. U.S.A.

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HPV-16 E6 and E7 genes are required to efficiently immortalize a broad spectrum of cell types including cervical keratinocytes. Therefore, the E6͞E7 genes can be considered relevant targets for anti-cancer therapy. We produced several engineered hairpin (HP) ribozymes to specifically disrupt HPV-16 E6͞E7 mRNA. After extensive biochemical characterization, one anti-E6 HP ribozyme (R434) was selected for in vivo testing because of its superior catalytic capabilities. When expressed in cis, R434 efficiently inhibited E6 in vitro translation. Cis-expression of the HP ribozyme with HPV-16 E6͞E7 genes in normal human keratinocytes reduced the growth rate and prevented immortalization. RNA analysis by reverse transcription-PCR showed that E6͞E7 transcripts were cleaved in post-transfected cells and virtually were eliminated after long term expression. Of interest, an inactive version of the HP also was able to significantly affect the immortalizing ability of E6͞E7, probably through passive hybridization. The combination of passive and cleaving antisense RNA therefore is established as an effective inhibitor of HPV-16 E6͞E7 immortalization.Over one-half of invasive cervical carcinomas worldwide and many cervical carcinoma-derived cell lines contain and express DNA from human papillomavirus type 16 (HPV-16). In early stages, HPV-16 expression causes benign proliferation and efficiently immortalizes cultured human epithelial cells, including cervical keratinocytes (1-3). The HPV-16 viral, early genes E6 and E7 are required to acquire and maintain efficiently the transformed phenotype in a broad spectrum of cell types and commonly are expressed in cervical carcinoma cells. Thus, E6͞E7 genes are the hallmark of cervical carcinoma (4-7). The E6 and E7 proteins bind with high affinity the p53 and Rb tumor suppressors, respectively (8, 9). The interaction of HPV-16 E6 protein with p53 results in degradation through the ubiquitin pathway, resulting in the equivalent of a mutant p53 phenotype (10-12). The association of E7 with Rb impedes the interaction of Rb with several proteins (i.e., E2F), efficiently disrupting the cell cycle (13-15). Therefore, the E6͞E7 genes are ideal targets for anti-cancer therapy.Antisense RNA and oligonucleotides have been used specifically to block translation of several genes. This effect is obtained by the hybridization of passive antisense molecules with their respective complementary mRNA to form nontranslatable double-stranded RNA molecules or DNA-RNA hybrids that promote the activity of endogenous RNase H, an enzyme that specifically digests the RNA strand of DNA-RNA hybrid molecules (16)(17)(18). In cultured tumor cells, antisense oligonucleotides have been shown to suppress effectively translation of several genes and to reverse some phenotypes (19)(20)(21)(22). However, passive antisense therapy has the disadvantage of being active for a limited period and often causing nonspecific toxicity (23,24). This approach has produced inhibition of E6͞E7 gene expression in HPV-18-containin...

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“…However, a recent report has shown that highrisk HPV E6 can degrade p53 protein even in the absence of E6AP ( . Low-risk HPV E6 oncoproteins can also bind to p53, but with very low affi nity and do not degrade p53 (Slebos et al 1995). There is accumulation of p53 after DNA damage in cells expressing HPV-11 E6 protein and the cells are arrested in the G1 phase of the cell cycle (Slebos et al 1995).…”

Section: Hpv E6 and E7 Oncoproteins And Cellular Transformationmentioning

confidence: 99%

“…Low-risk HPV E6 oncoproteins can also bind to p53, but with very low affi nity and do not degrade p53 (Slebos et al 1995). There is accumulation of p53 after DNA damage in cells expressing HPV-11 E6 protein and the cells are arrested in the G1 phase of the cell cycle (Slebos et al 1995). It has been further shown that low-risk HPV E6 proteins do not bind E6AP to form a trimeric complex of E6/E6AP/p53 like the highrisk E6 proteins (Zanier et al 2005).…”

Section: Hpv E6 and E7 Oncoproteins And Cellular Transformationmentioning

confidence: 99%

Human papillomavirus E6 and E7 oncoproteins as risk factors for tumorigenesis

2009

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Human papillomavirus (HPV) is small, double-stranded DNA virus that infects mucosal and cutaneous epithelial tissue. HPV is sexually transmitted and the viral DNA replicates extrachromosomally. The virus is non-enveloped and has an icosahedral capsid. There are approximately 118 types of HPV, which are characterized as high-risk or low- risk types. High-risk HPVs cause malignant transformation while the low-risk ones cause benign warts and lesions. The expression of E6 and E7 is normally controlled during the normal viral life cycle when viral DNA replicates extrachromosomally. HPV E6 and E7 oncoproteins are overexpressed when the viral genome integrates into the host DNA.Deregulated overexpression of E6 and E7 oncoproteins can cause several changes in cellular pathways and functions leading to malignant transformation of cells and tumorigenesis. In this review, we focus on several cellular mechanisms and pathways that are altered in the presence of E6 and E7, the target proteins of E6 and E7 inside the host cell and how they contribute to the development of the transformed phenotype.

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Functional Consequences of Directed Mutations in Human Papillomavirus E6 Proteins: Abrogation of p53-Mediated Cell Cycle Arrest Correlates with p53 Binding and Degradation in Vitro

Cited by 21 publications

References 0 publications

p53 Degradation Activity, Expression, and Subcellular Localization of E6 Proteins from 29 Human Papillomavirus Genotypes

p53 Degradation Activity, Expression, and Subcellular Localization of E6 Proteins from 29 Human Papillomavirus Genotypes

Inhibition of HPV-16 E6/E7 immortalization of normal keratinocytes by hairpin ribozymes

Human papillomavirus E6 and E7 oncoproteins as risk factors for tumorigenesis

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