scholarly journals The E6 Oncoproteins of High-Risk Papillomaviruses Bind to a Novel Putative GAP Protein, E6TP1, and Target It for Degradation

1999 ◽  
Vol 19 (1) ◽  
pp. 733-744 ◽  
Author(s):  
Qingshen Gao ◽  
Seetha Srinivasan ◽  
Sarah N. Boyer ◽  
David E. Wazer ◽  
Vimla Band
Keyword(s):  
High Risk ◽  
P53 Protein ◽  
Single Gene ◽  
Human Papillomaviruses ◽  
Dominant Negative ◽  
Hpv16 E6 ◽  
Hpv E6 ◽  
E6 Oncoprotein ◽  
E6 Protein

ABSTRACT The high-risk human papillomaviruses (HPVs) are associated with carcinomas of the cervix and other genital tumors. Previous studies have identified two viral oncoproteins, E6 and E7, which are expressed in the majority of HPV-associated carcinomas. The ability of high-risk HPV E6 protein to immortalize human mammary epithelial cells (MECs) has provided a single-gene model to study the mechanisms of E6-induced oncogenic transformation. In this system, the E6 protein targets the p53 tumor suppressor protein for degradation, and mutational analyses have shown that E6-induced degradation of p53 protein is required for MEC immortalization. However, the inability of most dominant-negative p53 mutants to induce efficient immortalization of MECs suggests the existence of additional targets of the HPV E6 oncoprotein. Using the yeast two-hybrid system, we have isolated a novel E6-binding protein. This polypeptide, designated E6TP1 (E6-targeted protein 1), exhibits high homology to GTPase-activating proteins for Rap, including SPA-1, tuberin, and Rap1GAP. The mRNA for E6TP1 is widely expressed in tissues and in vitro-cultured cell lines. The gene for E6TP1 localizes to chromosome 14q23.2-14q24.3 within a locus that has been shown to undergo loss of heterozygosity in malignant meningiomas. Importantly, E6TP1 is targeted for degradation by the high-risk but not the low-risk HPV E6 proteins both in vitro and in vivo. Furthermore, the immortalization-competent but not the immortalization-incompetent HPV16 E6 mutants target the E6TP1 protein for degradation. Our results identify a novel target for the E6 oncoprotein and provide a potential link between HPV E6 oncogenesis and alteration of a small G protein signaling pathway.

scholarly journals Cloning and expression of the cDNA for E6-AP, a protein that mediates the interaction of the human papillomavirus E6 oncoprotein with p53.

1993 ◽  
Vol 13 (2) ◽  
pp. 775-784 ◽  
Author(s):  
J M Huibregtse ◽  
M Scheffner ◽  
P M Howley
Keyword(s):  
High Risk ◽  
P53 Protein ◽  
Growth Control ◽  
Human Papillomaviruses ◽  
Cloning And Expression ◽  
Hpv16 E6 ◽  
Reading Frame ◽  
E6 Oncoprotein ◽  
Protein Complex Formation

The E6 oncoproteins of the cancer-associated or high-risk human papillomaviruses (HPVs) target the cellular p53 protein. The association of E6 with p53 leads to the specific ubiquitination and degradation of p53 in vitro, suggesting a model by which E6 deregulates cell growth control by the elimination of the p53 tumor suppressor protein. Complex formation between E6 and p53 requires an additional cellular factor, designated E6-AP (E6-associated protein), which has a native and subunit molecular mass of approximately 100 kDa. Here we report the purification of E6-AP and the cloning of its corresponding cDNA, which contains a novel open reading frame encoding 865 amino acids. E6-AP, translated in vitro, has the following properties: (i) it associates with wild-type p53 in the presence of the HPV16 E6 protein and simultaneously stimulates the association of E6 with p53, (ii) it associates with the high-risk HPV16 and HPV18 E6 proteins in the absence of p53, and (iii) it induces the E6- and ubiquitin-dependent degradation of p53 in vitro.

Cloning and expression of the cDNA for E6-AP, a protein that mediates the interaction of the human papillomavirus E6 oncoprotein with p53

1993 ◽  
Vol 13 (2) ◽  
pp. 775-784 ◽  
Author(s):  
J M Huibregtse ◽  
M Scheffner ◽  
P M Howley
Keyword(s):  
High Risk ◽  
P53 Protein ◽  
Growth Control ◽  
Human Papillomaviruses ◽  
Cloning And Expression ◽  
Hpv16 E6 ◽  
Reading Frame ◽  
E6 Oncoprotein ◽  
Protein Complex Formation

The E6 oncoproteins of the cancer-associated or high-risk human papillomaviruses (HPVs) target the cellular p53 protein. The association of E6 with p53 leads to the specific ubiquitination and degradation of p53 in vitro, suggesting a model by which E6 deregulates cell growth control by the elimination of the p53 tumor suppressor protein. Complex formation between E6 and p53 requires an additional cellular factor, designated E6-AP (E6-associated protein), which has a native and subunit molecular mass of approximately 100 kDa. Here we report the purification of E6-AP and the cloning of its corresponding cDNA, which contains a novel open reading frame encoding 865 amino acids. E6-AP, translated in vitro, has the following properties: (i) it associates with wild-type p53 in the presence of the HPV16 E6 protein and simultaneously stimulates the association of E6 with p53, (ii) it associates with the high-risk HPV16 and HPV18 E6 proteins in the absence of p53, and (iii) it induces the E6- and ubiquitin-dependent degradation of p53 in vitro.

scholarly journals Papillomavirus Type 16 Oncogenes Downregulate Expression of Interferon-Responsive Genes and Upregulate Proliferation-Associated and NF-κB-Responsive Genes in Cervical Keratinocytes

2001 ◽  
Vol 75 (9) ◽  
pp. 4283-4296 ◽  
Author(s):  
Matthias Nees ◽  
Joel M. Geoghegan ◽  
Tehila Hyman ◽  
Stephan Frank ◽  
Lance Miller ◽  
...  
Keyword(s):  
Cell Cycle Progression ◽  
P53 Protein ◽  
Human Papillomaviruses ◽  
Dominant Negative ◽  
Hpv 16 ◽  
Interacting Protein ◽  
Hpv E6 ◽  
E6 And E7 ◽  
E6 Protein ◽  
Functional Components

ABSTRACT Infection with high-risk human papillomaviruses (HPV) is a major risk factor for development of cervical cancer. Expression of the HPV E6 and E7 oncoproteins increases in differentiating keratinocytes, resulting in inactivation of the p53 and retinoblastoma proteins, two important transcriptional regulators. We used cDNA microarrays to examine global alterations in gene expression in differentiating cervical keratinocytes after infection with retroviruses encoding HPV type 16 (HPV-16) E6 and E7. Expression of 80 cellular genes (approximately 4% of the genes on the array) was altered reproducibly by E6 and/or E7. Cluster analysis classified these genes into three functional groups: (i) interferon (IFN)-responsive genes, (ii) genes stimulated by NF-κB, and (iii) genes regulated in cell cycle progression and DNA synthesis. HPV-16 E6 or a dominant negative p53 protein downregulated multiple IFN-responsive genes. E6 decreased expression of IFN-α and -β, downregulated nuclear STAT-1 protein, and decreased binding of STAT-1 to the IFN-stimulated response element. E7 alone was less effective; however, coexpression of E6 and E7 downregulated IFN-responsive genes more efficiently than E6. The HPV-16 E6 protein also stimulated expression of multiple genes known to be inducible by NF-κB and AP-1. E6 enhanced expression of functional components of the NF-κB signal pathway, including p50, NIK, and TRAF-interacting protein, and increased binding to NF-κB and AP-1 DNA consensus binding sites. Secretion of interleukin-8, RANTES, macrophage inflammatory protein 1α, and 10-κDa IFN-γ-inducible protein were increased in differentiating keratinocytes by E6. Thus, high-level expression of the HPV-16 E6 protein in differentiating keratinocytes directly alters expression of genes that influence host resistance to infection and immune function.

scholarly journals High Risk α-HPV E6 Impairs Translesion Synthesis by Blocking POLη Induction

Cancers ◽  
2020 ◽  
Vol 13 (1) ◽  
pp. 28
Author(s):  
Sebastian O. Wendel ◽  
Jazmine A. Snow ◽  
Tyler Bastian ◽  
Laura Brown ◽  
Candy Hernandez ◽  
...  
Keyword(s):  
Dna Damage ◽  
High Risk ◽  
Replication Fork ◽  
Ex Vivo ◽  
Translesion Synthesis ◽  
Replication Stress ◽  
Human Papillomaviruses ◽  
Hpv16 E6 ◽  
Hpv E6

High risk genus α human papillomaviruses (α-HPVs) express two versatile oncogenes (α-HPV E6 and E7) that cause cervical cancer (CaCx) by degrading tumor suppressor proteins (p53 and RB). α-HPV E7 also promotes replication stress and alters DNA damage responses (DDR). The translesion synthesis pathway (TLS) mitigates DNA damage by preventing replication stress from causing replication fork collapse. Computational analysis of gene expression in CaCx transcriptomic datasets identified a frequent increased expression of TLS genes. However, the essential TLS polymerases did not follow this pattern. These data were confirmed with in vitro and ex vivo systems. Further interrogation of TLS, using POLη as a representative TLS polymerase, demonstrated that α-HPV16 E6 blocks TLS polymerase induction by degrading p53. This doomed the pathway, leading to increased replication fork collapse and sensitivity to treatments that cause replication stress (e.g., UV and Cisplatin). This sensitivity could be overcome by the addition of exogenous POLη.

scholarly journals The Human Papillomavirus (HPV) E6* Proteins from High-Risk, Mucosal HPVs Can Direct Degradation of Cellular Proteins in the Absence of Full-Length E6 Protein

2009 ◽  
Vol 83 (19) ◽  
pp. 9863-9874 ◽  
Author(s):  
David Pim ◽  
Vjekoslav Tomaić ◽  
Lawrence Banks
Keyword(s):  
High Risk ◽  
Tumor Suppressor ◽  
Pdz Domain ◽  
Direct Interaction ◽  
Human Papillomaviruses ◽  
Full Length ◽  
Cellular Proteins ◽  
Hpv E6 ◽  
E6 Protein ◽  
Hpv 18

ABSTRACT The E6 oncoproteins from high-risk mucosotrophic human papillomaviruses (HPVs) target a range of cellular proteins for proteasome-mediated degradation. Apart from the tumor suppressor p53 and proapoptotic Bcl-2 family member Bak, many targets contain class 1 PDZ domains and are involved in cell junction stability and signaling. The targeting mechanism is considered to function by the E6 protein acting as an adaptor molecule linking a cellular ubiquitin ligase to the target protein. In each case, whether the target is the p53 tumor suppressor or a member of the group of PDZ domain-containing targets, this mechanism relies on a direct interaction between E6 and its cellular target. This study focuses on the impact of the HPV type 18 (HPV-18) E6*I protein on the stability of Akt, Dlg, MAGI-1, MAGI-2, and Scribble. We show that HPV-18 E6* expression can downregulate the expression levels of Akt, Dlg, and Scribble in the absence of full-length HPV-18 E6 protein. The reduction in Dlg levels by E6* is independent of transcription and does not require a direct interaction between the two proteins although the proteasome pathway is involved. Further, we provide evidence that activation of certain signal transduction pathways has a profound effect on the targeting of Dlg by E6* and suggest that high-risk HPV E6 oncoproteins can target certain substrates both directly and indirectly through the E6* proteins and may cooperate in their degradation.

scholarly journals Human Papillomavirus Oncoprotein E6 Inactivates the Transcriptional Coactivator Human ADA3

2002 ◽  
Vol 22 (16) ◽  
pp. 5801-5812 ◽  
Author(s):  
Ajay Kumar ◽  
Yongtong Zhao ◽  
Gaoyuan Meng ◽  
Musheng Zeng ◽  
Seetha Srinivasan ◽  
...  
Keyword(s):  
High Risk ◽  
P53 Protein ◽  
Human Papillomaviruses ◽  
Transcriptional Coactivator ◽  
Interacting Protein ◽  
Hpv E6 ◽  
Cycle Arrest ◽  
Evolutionarily Conserved ◽  
Novel Strategy ◽  
High Risk Hpv

ABSTRACT High-risk human papillomaviruses (HPVs) are associated with carcinomas of the cervix and other genital tumors. The HPV oncoprotein E6 is essential for oncogenic transformation. We identify here hADA3, human homologue of the yeast transcriptional coactivator yADA3, as a novel E6-interacting protein and a target of E6-induced degradation. hADA3 binds selectively to the high-risk HPV E6 proteins and only to immortalization-competent E6 mutants. hADA3 functions as a coactivator for p53-mediated transactivation by stabilizing p53 protein. Notably, three immortalizing E6 mutants that do not induce direct p53 degradation but do interact with hADA3 induced the abrogation of p53-mediated transactivation and G1 cell cycle arrest after DNA damage, comparable to wild-type E6. These findings reveal a novel strategy of HPV E6-induced loss of p53 function that is independent of direct p53 degradation. Given the likely role of the evolutionarily conserved hADA3 in multiple coactivator complexes, inactivation of its function may allow E6 to perturb numerous cellular pathways during HPV oncogenesis.

scholarly journals Transforming Properties of Beta-3 Human Papillomavirus E6 and E7 Proteins

mSphere ◽  
2020 ◽  
Vol 5 (4) ◽  
Author(s):  
Lucia Minoni ◽  
Maria Carmen Romero-Medina ◽  
Assunta Venuti ◽  
Cécilia Sirand ◽  
Alexis Robitaille ◽  
...  
Keyword(s):  
High Risk ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Biological Properties ◽  
Human Papillomaviruses ◽  
Hpv E6 ◽  
Hpv Types ◽  
Beta 2 ◽  
E6 And E7

ABSTRACT The beta human papillomaviruses (HPVs) are subdivided into 5 species (beta-1 to beta-5), and they were first identified in the skin. However, the beta-3 species appears to be more highly represented in the mucosal epithelia than in the skin. Functional studies have also highlighted that beta-3 HPV49 shares some functional similarities with mucosal high-risk (HR) HPV16. Here, we describe the characterization of the in vitro transforming properties of the entire beta-3 species, which includes three additional HPV types: HPV75, HPV76, and HPV115. HPV49, HPV75, and HPV76 E6 and E7 (E6/E7), but not HPV115 E6 and E7, efficiently inactivate the p53 and pRb pathways and immortalize or extend the life span of human foreskin keratinocytes (HFKs). As observed for HR HPV16, cell cycle deregulation mediated by beta-3 HPV E6/E7 expression leads to p16INK4a accumulation, whereas no p16INK4a was detected in beta-2 HPV38 E6/E7 HFKs. As shown for HPV49 E6, HPV75 and HPV76 E6s degrade p53 by an E6AP/proteasome-mediated mechanism. Comparative analysis of cellular gene expression patterns of HFKs containing E6 and E7 from HR HPV16, beta-3 HPV types, and beta-2 HPV38 further highlights the functional similarities of HR HPV16 and beta-3 HPV49, HPV75, and HPV76. The expression profiles of these four HPV HFKs show some similarities and diverge substantially from those of beta-3 HPV115 E6/E7 and beta-2 HPV38 E6/E7 HFKs. In summary, our data show that beta-3 HPV types share some mechanisms with HR HPV types and pave the way for additional studies aiming to evaluate their potential role in human pathologies. IMPORTANCE Human papillomaviruses are currently classified in different genera. Mucosal HPVs belonging to the alpha genus have been clearly associated with carcinogenesis of the mucosal epithelium at different sites. Beta HPV types have been classified as cutaneous. Although findings indicate that some beta HPVs from species 1 and 2 play a role, together with UV irradiation, in skin cancer, very little is known about the transforming properties of most of the beta HPVs. This report shows the transforming activity of E6 and E7 from beta-3 HPV types. Moreover, it highlights that beta-3 HPVs share some biological properties more extensively with mucosal high-risk HPV16 than with beta-2 HPV38. This report provides new paradigms for a better understanding of the biology of the different HPV types and their possible association with lesions at mucosal and/or cutaneous epithelia.

scholarly journals Transactivation activity of human papillomavirus type 16 E6*I on aldo-keto reductase genes enhances chemoresistance in cervical cancer cells

2012 ◽  
Vol 93 (5) ◽  
pp. 1081-1092 ◽  
Author(s):  
Panata Wanichwatanadecha ◽  
Sasinan Sirisrimangkorn ◽  
Jittranan Kaewprag ◽  
Mathurose Ponglikitmongkol
Keyword(s):  
Cervical Cancer ◽  
Drug Resistance ◽  
High Risk ◽  
Intracellular Localization ◽  
Human Papillomaviruses ◽  
Moderate Increase ◽  
Transcript Levels ◽  
Hpv16 E6 ◽  
Transactivation Activity ◽  
E6 Protein

The oncogenic E6 proteins produced by high-risk human papillomaviruses (HPVs) are invariably expressed in cervical carcinomas and are multifunctional proteins capable of affecting host-cell proliferation by binding and deregulating key host molecules such as p53. High-risk HPVs, including HPV16, have the unique ability to splice the E6 viral transcript, resulting in the production of a truncated E6 protein known as E6*I whose precise biological function is unclear. This study explored the changes in gene expression of the cervical cancer C33A cell line stably expressing HPV16 E6*I (16E6*I) and observed the upregulation of ten genes. Two of these genes were aldo-keto reductases (AKR1Cs), AKR1C1 and AKR1C3, which have been implicated in drug resistance. The results demonstrated that expression of 16E6*I, but not full-length E6, specifically increased AKR1C1 transcript levels although it did not alter AKR1C2 transcript levels. HPV16 E7 alone also had the ability to cause a moderate increase in AKR1C3 at both mRNA and protein levels. Site-directed mutagenesis of 16E6*I revealed that transactivation activity was abolished in R8A, R10A and T17A 16E6*I mutants without altering their intracellular localization patterns. Loss of transactivation activity of the 16E6*I mutants resulted in a significant loss of AKR1C expression and a decrease in drug resistance. Analysis of the AKR1C1 promoter revealed that, unlike the E6 protein, 16E6*I does not mediate transactivation activity solely through Sp1-binding sites. Taken together, it was concluded that 16E6*I has a novel function in upregulating expression of AKR1C and, in concert with E7, has implications for drug treatment in HPV-mediated cervical cancer.

scholarly journals Cellular Distribution of Tumour Suppressor Protein p53 and High-Risk Human Papillomavirus (HPV)-18 E6 Fusion Protein in Wild-Type p53 Cell Lines

2008 ◽  
Vol 36 (5) ◽  
pp. 1015-1021 ◽  
Author(s):  
L Sun ◽  
G Gzhang ◽  
Z Li ◽  
T Lei ◽  
C Huang ◽  
...  
Keyword(s):  
High Risk ◽  
Fluorescent Protein ◽  
P53 Protein ◽  
Expression System ◽  
Human Papillomaviruses ◽  
Tumour Suppressor ◽  
Wild Type ◽  
Hpv E6 ◽  
Wild Type P53 ◽  
High Risk Hpv

Human papillomaviruses (HPVs) are very important pathogens that can be classified as high- and low-risk types based on the lesions they cause. Mucosal high-risk HPV E6 can target and degrade the tumour suppressor p53, hence it is recognized as the major cause of cervical cancer, however, due to a lack of reliable anti-E6 antibodies, the distribution of high-risk HPV E6 protein remains elusive. The present study, therefore, used a mammalian green fluorescent protein (GFP) expression system to express GFP-18 E6 fusion proteins in wild-type p53 cells, SMMC-7721 and HCT116, in order to trace the location and expression of HPV E6 and p53 protein. Following transfection, expression of GFP-18 E6 was found to be located in the nucleus, and endogenous wild-type p53 was also located there with GFP-18 E6.

scholarly journals Role of ubiquitin and the HPV E6 oncoprotein in E6AP-mediated ubiquitination

2015 ◽  
Vol 112 (32) ◽  
pp. 9872-9877 ◽  
Author(s):  
Franziska Mortensen ◽  
Daniel Schneider ◽  
Tanja Barbic ◽  
Anna Sladewska-Marquardt ◽  
Simone Kühnle ◽  
...  
Keyword(s):  
Noncovalent Interactions ◽  
Neurodevelopmental Disorder ◽  
Autism Spectrum ◽  
Human Papillomaviruses ◽  
Hydrophobic Patch ◽  
Hpv E6 ◽  
E6 Oncoprotein ◽  
Clinical Pictures

Deregulation of the ubiquitin ligase E6 associated protein (E6AP) encoded by the UBE3A gene has been associated with three different clinical pictures. Hijacking of E6AP by the E6 oncoprotein of distinct human papillomaviruses (HPV) contributes to the development of cervical cancer, whereas loss of E6AP expression or function is the cause of Angelman syndrome, a neurodevelopmental disorder, and increased expression of E6AP has been involved in autism spectrum disorders. Although these observations indicate that the activity of E6AP has to be tightly controlled, only little is known about how E6AP is regulated at the posttranslational level. Here, we provide evidence that the hydrophobic patch of ubiquitin comprising Leu-8 and Ile-44 is important for E6AP-mediated ubiquitination, whereas it does not affect the catalytic properties of the isolated catalytic HECT domain of E6AP. Furthermore, we show that the HPV E6 oncoprotein rescues the disability of full-length E6AP to use a respective hydrophobic patch mutant of ubiquitin for ubiquitination and that it stimulates E6AP-mediated ubiquitination of Ring1B, a known substrate of E6AP, in vitro and in cells. Based on these data, we propose that E6AP exists in at least two different states, an active and a less active or latent one, and that the activity of E6AP is controlled by noncovalent interactions with ubiquitin and allosteric activators such as the HPV E6 oncoprotein.

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