Binding of PDZ proteins to HPV E6 proteins does neither correlate with epidemiological risk classification nor with the immortalization of foreskin keratinocytes

Muench, Peter; Hiller, Thomas; Probst, Sonja; Florea, Ana-Maria; Stubenrauch, Frank; Iftner, Thomas

doi:10.1016/j.virol.2009.02.018

Cited by 30 publications

(40 citation statements)

References 40 publications

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“…These findings are in accordance with earlier studies showing successful immortalization of primary keratinocytes by full-length HPV66 and HPV70 E6/E7 (11,14). This study, however, is the first to systematically demonstrate that these probable hrHPV types display a reduced immortalization capacity compared to the established hrHPV types 16, 18, 31, 33, and 35.…”

Section: Discussionsupporting

confidence: 92%

“…Furthermore, a large, randomized, controlled prospective trial conducted in the Netherlands has revealed that among hrHPV-positive women with normal cytology or borderline to moderate dyskaryosis, those infected with hrHPV types 16, 18, 31, and 33 have an increased 18-month risk of CIN3 or cancer (8). Apart from type-specific differences in viral persistence (9), these and other data also point to the existence of different oncogenic properties of the various high-risk types.Although some non-HPV16 and -18 types recently received some attention (11)(12)(13)(14), functional studies on transforming properties of hrHPV are mainly limited to HPV16 and HPV18. We and others have previously demonstrated that HPV16 and HPV18 can induce immortalization of primary human foreskin keratinocytes (HFKs) by successive bypass of two proliferative life span barriers, i.e., senescence and crisis (reviewed in reference 15).…”

mentioning

confidence: 99%

“…Although some non-HPV16 and -18 types recently received some attention (11)(12)(13)(14), functional studies on transforming properties of hrHPV are mainly limited to HPV16 and HPV18. We and others have previously demonstrated that HPV16 and HPV18 can induce immortalization of primary human foreskin keratinocytes (HFKs) by successive bypass of two proliferative life span barriers, i.e., senescence and crisis (reviewed in reference 15).…”

mentioning

confidence: 99%

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Differential In Vitro Immortalization Capacity of Eleven, Probable High-Risk Human Papillomavirus Types

Schütze

Snijders

Bosch

et al. 2014

J Virol

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bEpidemiological studies identified 12 high-risk HPV (hrHPV) types and 8 probable/possible hrHPV types that display different cancer risks. Functional studies on transforming properties of hrHPV are mainly limited to HPV16 and -18, which induce immortalization of human foreskin keratinocytes (HFKs) by successive bypass of two proliferative life span barriers, senescence and crisis. Here, we systematically compared the in vitro immortalization capacities, as well as influences on p53, pRb, hTERT, growth behavior, and differentiation capacity, of nine hrHPV types (HPV16, -18, -31, -33, -35, -45, -51, -52, and -59), and two probable hrHPV types (HPV66 and -70). By retroviral transduction, the respective E6/E7 coding sequences were expressed in HFKs from two or three independent donors. Reduced p53 levels and low-level hTERT expression in early-passage cells, as seen in HPV16-, -31-, -33-, and -35-, and to a lesser extent HPV18-transduced HFKs, was associated with continuous growth and an increased immortalization capacity. Less frequent immortalization by HPV45 and -51 and immortalization by HPV66 and -70 was preceded by an intervening period of strongly reduced growth (crisis) without prior increase in hTERT expression. Immortalization by HPV59 was also preceded by a period crisis, despite the onset of low hTERT expression at early passage. HPV52 triggered an extended life span but failed to induce immortality. Variations in p53 and pRb levels were not correlated with differences in alternative E6/E7 mRNA splicing in all hrHPV-transduced HFKs. On collagen rafts, transductants showed disturbed differentiation reminiscent of precancerous lesions. In conclusion, in vitro oncogenic capacities differ between the established hrHPV types, and both some established and probable hrHPV types display weak or moderate immortalization potential. C ervical cancer, the third most common cancer among women worldwide, is caused by a persistent infection with certain types of human papillomavirus (HPV) (1, 2). Most HPV infections are transient and are cleared within 1 to 2 years. However, a fraction of infections eventually give rise to either squamous cell carcinoma (SCC) or adenocarcinoma (AdCA) of the cervix. SCCs develop from so-called cervical intraepithelial neoplasia (CIN) precursor lesions. Low-grade CIN lesions (CIN1) mostly reflect a productive infection in which viral replication and virion production are linked to the differentiation program of the epithelium. High-grade CIN lesions (CIN2/3) mainly represent transforming infections and are characterized by deregulated expression of the early viral E6 and E7 genes in the proliferating basal cells of the epithelium (3, 4).HPV types belonging to the alpha (␣) genus can infect the cervical mucosa (5) and are classified into low-risk (lrHPV) and high-risk (hrHPV) HPV based on their association with malignancy (6). Infections with lrHPV types, e.g., HPV type 6 (HPV6) and HPV11, are associated with benign warts or low-grade lesions, whereas infections with hrHPV can give rise ...

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Section: Discussionsupporting

confidence: 92%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Differential In Vitro Immortalization Capacity of Eleven, Probable High-Risk Human Papillomavirus Types

Schütze

Snijders

Bosch

et al. 2014

J Virol

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“…Only a limited number of studies have addressed the transforming ability of the beta HPV types in primary keratinocytes (6,9,15). In the present work, we compared the transforming potential of E6 and E7 proteins from several HPV types, which belong to different subgroups within the beta genus, i.e., HPV14, -24, and -36 (␤1), HPV-22, -23, and -38 (␤2), and HPV49 (␤3).…”

mentioning

confidence: 99%

Comparative Analysis of Transforming Properties of E6 and E7 from Different Beta Human Papillomavirus Types

Cornet¹,

Bouvard²,

Campo³

et al. 2012

J Virol

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The cutaneous beta human papillomavirus (beta HPV) types appear to be involved in skin carcinogenesis. However, only a few beta HPVs have been investigated so far. Here, we compared the properties of E6 and E7 oncoproteins from six uncharacterized beta HPVs (14, 22, 23, 24, 36, 49). Only HPV49 E6 and E7 immortalized primary human keratinocytes and efficiently deregulated the p53 and pRb pathways. Furthermore, HPV49 E6, similarly to E6 from the oncogenic HPV16, promoted p53 degradation. The cutaneous human papillomavirus (HPV) types that belong to the beta genus are suspected to play a role in skin carcinogenesis together with UV irradiation (4, 17). They were originally isolated from patients suffering from a rare autosomal recessive genetic disorder called epidermodysplasia verruciformis (EV), who have an increased susceptibility to beta HPV infection and who develop nonmelanoma skin cancer (NMSC) (17). Epidemiological studies have also provided evidence for the role of beta HPV types in NMSC in immunocompromised individuals, such as organ transplant recipients, as well as in the normal population (2,3,5,7,8,(12)(13)(14)18). However, due to the high frequency with which cutaneous HPV DNA is found in the skin of healthy individuals, the role of these viruses in carcinogenesis is still unclear.Studies on the cervical cancer-associated HPV types (for example, see references 16 and 18) showed that their oncogenic potential lies in the transforming activities of two proteins, E6 and E7 (11). By interacting with several host proteins, E6 and E7 of highrisk (HR) HPV types are able to interfere with key cellular processes, such as cell cycle, senescence, differentiation, apoptosis, and telomere shortening.Only a limited number of studies have addressed the transforming ability of the beta HPV types in primary keratinocytes (6,9,15). In the present work, we compared the transforming potential of E6 and E7 proteins from several HPV types, which belong to different subgroups within the beta genus, i.e., HPV14, -24, and -36 (␤1), HPV-22, -23, and -38 (␤2), and HPV49 (␤3).Beta HPV49 E6 and E7 efficiently immortalize primary human keratinocytes. We first compared the ability of E6 and E7 from different beta HPV types to immortalize primary cells. Primary human foreskin keratinocytes (HFK) were transduced with E6/E7 recombinant retroviruses, and viral gene expression was determined by reverse transcription-PCR (RT-PCR) using specific primers for the different E7 genes (10) (Fig. 1A). Twenty days after retroviral transduction, HPV14 and -22 E6/E7-transduced HFK displayed features of senescence, such as irregular shape and intercellular bridges, similar to the HFK transduced with the empty retroviral vector (pLXSN) (mock-transduced HFK) (Fig. 1B). Consistently, all three HFK cultures showed high positivity for the senescent marker ␤-galactosidase (data not shown) and died after a few population doublings (PD) ( Table 1). HPV23, -24, -36, and -38 E6/E7 HFK continued to grow for a few PDs after transduction, then stopped prolife...

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“…The generation of amphotropic recombinant retroviruses and infection of NHK was performed as previously described (37). Transduced cells were selected with puromycin (0.4 μg/mL) or G418 (50 μg/mL) for 14 days.…”

Section: Generation and Analysis Of Stable Keratinocyte Cell Linesmentioning

confidence: 99%

Cutaneous Papillomavirus E6 Proteins Must Interact with p300 and Block p53-Mediated Apoptosis for Cellular Immortalization and Tumorigenesis

Muench¹,

Probst²,

Schuetz³

et al. 2010

Cancer Research

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The binding of the papillomavirus E6 protein to E6AP and the induction of p53 degradation are common features of high-risk genital human papillomaviruses (HPV); cutaneous HPVs, on the other hand, lack these capacities. Nevertheless, several cutaneous HPV types of the β-genus, such as HPV38 are associated with tumor formation when combined with genetic predisposition, immunosuppression, or UV exposure. In an animal model system, the cottontail rabbit papillomavirus (CRPV) rapidly induces skin cancer without additional cofactors, and CRPVE6 and E7 immortalize rabbit keratinocytes in vitro. However, CRPVE6 neither interacts with E6AP and p53 nor does it induce p53 degradation. In this study, we show that the interaction of CRPVE6, or HPV38E6, with the histone acetyltransferase p300 is crucial to inhibit the ability of p53 to induce apoptosis. Strikingly, E6 mutants deficient for p300 binding are incapable of preventing p53 acetylation, p53-dependent transcription, and apoptosis induction. Moreover, E6 mutants deficient for p300 binding cannot contribute to HPV38-induced immortalization of human keratinocytes or CRPV-induced tumor formation. Our findings highlight changes in the p53 acetylation status mediated by the viral E6 protein as a crucial requirement in the ability of high-risk cutaneous papillomaviruses to immortalize primary keratinocytes and induce tumors.

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Binding of PDZ proteins to HPV E6 proteins does neither correlate with epidemiological risk classification nor with the immortalization of foreskin keratinocytes

Cited by 30 publications

References 40 publications

Differential In Vitro Immortalization Capacity of Eleven, Probable High-Risk Human Papillomavirus Types

Differential In Vitro Immortalization Capacity of Eleven, Probable High-Risk Human Papillomavirus Types

Comparative Analysis of Transforming Properties of E6 and E7 from Different Beta Human Papillomavirus Types

Cutaneous Papillomavirus E6 Proteins Must Interact with p300 and Block p53-Mediated Apoptosis for Cellular Immortalization and Tumorigenesis

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