Growth Inhibition of HeLa Cells Is a Conserved Feature of High-Risk Human Papillomavirus E8^E2C Proteins and Can Also Be Achieved by an Artificial Repressor Protein

Fertey, Jasmin; Hurst, José; Straub, Elke; Schenker, Astrid; Iftner, Thomas; Stubenrauch, Frank

doi:10.1128/jvi.01647-10

Cited by 20 publications

(34 citation statements)

References 46 publications

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“…This experiment was repeated three times with similar outcomes. These findings are similar to the effects reported by the reduction of E6 and E7 transcript levels in cervical cancer cell lines by siRNA or expression of E2 or E8^E2C proteins (18,(37)(38)(39)(40) and suggested that the decrease of viral genome copies and transcription levels resulted in lowered levels of the E6 and E7 oncoproteins, which causes a reduced proliferation.…”

Section: Resultssupporting

confidence: 77%

“…The pInducer 20-IFN-plasmid was constructed by in vitro recombination of pInducer20 (17) with pENTR4-IFN-. Reporter plasmids pGL18 URR and pGL31 URR have been previously described (18). Expression plasmids for SP100 isoforms and the shSp100 expression construct and the respective control were kindly provided by S. Schreiner and have been previously described (19)(20)(21).…”

Section: Methodsmentioning

confidence: 99%

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Interferon Kappa Inhibits Human Papillomavirus 31 Transcription by Inducing Sp100 Proteins

Habiger

Jäger²,

Walter³

et al. 2016

J Virol

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High-risk human papillomaviruses (hr-HPV IMPORTANCEHigh-risk HPV can establish persistent infections which may progress to anogenital cancers. hr-HPV interfere with the expression of interferon (IFN)-stimulated genes (ISGs), which is due to reduced levels of IFN-, an IFN that is constitutively expressed in human keratinocytes. This study reveals that IFN-rapidly inhibits HPV transcription and that this is due to the induction of Sp100 proteins. Thus, Sp100 represents an ISG for hr-HPV. P ersistent infections with high-risk human papillomaviruses (hr-HPV) are a necessary risk factor for the development of anogenital and oropharyngeal cancers (1). HPV have circular double-stranded DNA (dsDNA) genomes of ϳ8,000 bp and infect keratinocytes of the basal layer of mucosal and cutaneous epithelium. In undifferentiated cells, HPV genomes replicate as extrachromosomal elements at a low copy number and express only early viral genes such as those for the oncoproteins E6 and E7 and the replication proteins E1, E2, and E8^E2C (2). hr-HPV E6 and E7 interact with critical regulators of the cell cycle such as p53 and retinoblastoma family members to ensure continuous proliferation of infected cells (3).Transcriptome studies have shown that interferon (IFN)-stimulated genes (ISGs) are expressed at lower levels in hr-HPV-positive cell lines than in their uninfected counterparts (4-6). Type I IFNs such as IFN-␣ subtypes or IFN-␤ are induced upon viral infection and then secreted into the extracellular space (7). Secreted IFNs bind to the respective receptor on infected and neighboring cells, and kinases are activated, which results in the nuclear translocation of transcription factors such as STAT1 which then induce several hundred ISGs, many of which have direct antiviral activities (8).The treatment of cell lines that maintain persistently replicating extrachromosomal HPV16 or -31 genomes with recombinant IFN-␤ resulted in growth retardation and the induction of apoptosis, which did not occur with HPV-negative keratinocytes (9, 10). A detailed analysis of the HPV16 copy number and transcription upon IFN-␤ treatment indicated that first the viral copy number is decreased, followed by a reduction of viral transcription (10). HPV31-positive CIN612-9E cells have reduced levels of STAT1, which is both an ISG and also a crucial transcription factor of the IFN signal transduction cascade. Reexpression of STAT1 resulted in a reduction of viral genomes (11). Taken together, these data strongly suggest that IFNs induce ISGs that inhibit the replication of HPV. In line with this, the ISG IFIT1 (or ISG56) directly interacts with the E1 helicases of HPV11 and -18, which results in the inhibition of the replication of the viral origin (12).

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Section: Resultssupporting

confidence: 77%

Section: Methodsmentioning

confidence: 99%

Interferon Kappa Inhibits Human Papillomavirus 31 Transcription by Inducing Sp100 Proteins

Habiger

Jäger²,

Walter³

et al. 2016

J Virol

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“…In the E8 KWK mt, E8 residues K5/W6/K7 were changed to AAA, which also changes the E1 coding sequence from E138/V139/E140 to GSG. The pGL 16URR-luc and pC18-Sp1-luc luciferase reporter plasmids and the expression plasmids for HPV16 E2 and HPV16 E8^E2C and the respective hemagglutinin (HA)-tagged versions (HPV16 E2-HA and HPV16 E8^E2C-HA) have been previously described (27). Plasmid pSG 16 E8^E2C KWK mt and 16 E8^E2C-KWK mt -HA were constructed by PCR.…”

Section: Methodsmentioning

confidence: 99%

“…Genetic analyses have revealed that E8^E2C is a potent inhibitor of the genome replication of HR-HPV16, -18 and -31 in undifferentiated cells (29,30,34,35). HR-HPV E8^E2C proteins are efficient repressors of the major viral early promoter (27,30,34,36,37). In addition, E8^E2C proteins inhibit the E1-and E2-dependent replication of the viral origin (30,34,35).…”

mentioning

confidence: 99%

“…This transcript encodes an E8^E2C (or E8/E2) protein which consists of the E8 gene product fused to the C-terminal half of E2, which mediates sequence-specific DNA binding and dimerization of E2 proteins (16,(25)(26)(27)(28)(29)(30)(31)(32)(33)(34). Genetic analyses have revealed that E8^E2C is a potent inhibitor of the genome replication of HR-HPV16, -18 and -31 in undifferentiated cells (29,30,34,35).…”

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The Viral E8^E2C Repressor Limits Productive Replication of Human Papillomavirus 16

Straub

Dreer

Fertey

et al. 2014

J Virol

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Productive replication of human papillomavirus type 16 (HPV16) occurs only in differentiated keratinocyte cells. In addition to the viral E2 activator protein, HPV16 and related HPV types express transcripts coding for an E8^E2C fusion protein, which limits genome replication in undifferentiated keratinocytes. To address E8^E2C's role in productive replication of HPV16, stable keratinocyte cell lines containing wild-type (wt), E8^E2C knockout (E8؊), or E8 KWK mutant (mt) genomes, in which conserved E8 residues were inactivated, were established. Copy numbers of E8؊ and E8 KWK mt genomes and amounts of early and late viral transcripts were greatly increased compared to those for the wt in undifferentiated keratinocytes, suggesting that HPV16 E8^E2C activities are highly dependent upon the E8 part. Upon differentiation in organotypic cultures, E8 mt genomes displayed higher early viral transcript levels, but no changes in cellular differentiation or virus-induced cellular DNA replication in suprabasal cells were observed. E8 mt genomes were amplified to higher copy numbers and showed increased L1 transcripts compared to wt genomes. Furthermore, the number of cells expressing the viral late protein E4 or L1 or amplifying viral genomes was greatly increased in E8 mt cell lines. In wild-type cells, E8^E2C transcript levels did not decrease by differentiation. Our data indicate that the E8^E2C repressor limits viral transcription and replication throughout the complete life cycle of HPV16. P ersistent infections with high-risk (HR) human papillomaviruses (HPV) are a necessary risk factor for the development of cancer of the cervix uteri and are also responsible for a fraction of anal, vaginal, penile, and oropharyngeal cancers (1-3). Notably, HPV16 is an extremely powerful human carcinogen that is responsible for approximately 50% of cervical cancers and is also represented in other HR-HPV-related cancers at high frequencies (2).HPV have a double-stranded DNA genome that is covalently closed and organized in nucleosomes. HPV possess eight to nine open reading frames (E1, E2, E4 to E8, L1, and L2) that are transcribed into polycistronic messages which are further processed by alternative splicing (4). It is currently assumed that HR-HPV encode at least 10 different viral proteins.The HR-HPV replication cycle is tightly linked to the differentiation state of the infected epithelium. In undifferentiated basalcell-like cells, HR-HPV genomes are maintained in the nucleus at 10 to 100 extrachromosomal copies per cell, and only early genes, mainly transcribed from the major early viral promoter located immediately upstream of the E6 gene (P97 for HPV16), but not the late viral L1 and L2 genes are expressed (5). Replication of the viral genome in undifferentiated cells requires the viral E1 and E2 proteins (6). E1 and E2 are sequence-specific DNA binding proteins which form a complex that recognizes the viral origin of replication (6). E1 assembles into a hexamer that acts as the replicative helicase and unwinds the viral DNA...

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Epidemiology of HPV in Head and Neck Cancer: Variant Strains, Discrete Protein Function

Ragin

Liu

2014

Molecular Determinants of Head and Neck Cancer

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Growth Inhibition of HeLa Cells Is a Conserved Feature of High-Risk Human Papillomavirus E8^E2C Proteins and Can Also Be Achieved by an Artificial Repressor Protein

Cited by 20 publications

References 46 publications

Interferon Kappa Inhibits Human Papillomavirus 31 Transcription by Inducing Sp100 Proteins

Interferon Kappa Inhibits Human Papillomavirus 31 Transcription by Inducing Sp100 Proteins

The Viral E8^E2C Repressor Limits Productive Replication of Human Papillomavirus 16

Epidemiology of HPV in Head and Neck Cancer: Variant Strains, Discrete Protein Function

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