RNA Interference of Human Papillomavirus Type 18 E6 and E7 Induces Senescence in HeLa Cells

Hall, Allison; Alexander, Kenneth A.

doi:10.1128/jvi.77.10.6066-6069.2003

Cited by 184 publications

(174 citation statements)

References 17 publications

(19 reference statements)

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“…Knock down of E6 and E7 protein by RNA interference could induce senescence in HeLa Cells 30. Consistently, we observed that HeLa cells occurred senescence when the integrated HPV fragment was removed.…”

Section: Discussionsupporting

confidence: 83%

Long-distance interaction of the integrated HPV fragment with MYC gene and 8q24.22 region upregulating the allele-specific MYC expression in HeLa cells

et al. 2017

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Human papillomavirus (HPV) infection is the most important risk factor for cervical cancer development. In HeLa cell line, the HPV viral genome is integrated at 8q24 in one allele of chromosome 8. It has been reported that the HPV fragment integrated in HeLa genome can cis‐activate the expression of proto‐oncogene MYC, which is located at 500 kb downstream of the integrated site. However, the underlying molecular mechanism of this regulation is unknown. A recent study reported that MYC was highly expressed exclusively from the HPV‐integrated haplotype, and a long‐range chromatin interaction between the integrated HPV fragment and MYC gene has been hypothesized. In this study, we provided the experimental evidences supporting this long‐range chromatin interaction in HeLa cells by using Chromosome Conformation Capture (3C) method. We found that the integrated HPV fragment, MYC and 8q24.22 was close to each other and might form a trimer in spatial location. When knocking out the integrated HPV fragment or 8q24.22 region from chromosome 8 by CRISPR/Cas9 system, the expression of MYC reduced dramatically in HeLa cells. Interestingly, decreased expression was only observed in three from eight cell clones, when only one 8q24.22 allele was knocked out. Functionally, HPV knockout caused senescence‐associated acidic β‐gal activity in HeLa cells. These data indicate a long‐distance interaction of the integrated HPV fragment with MYC gene and 8q24.22 region, providing an alternative mechanism relevant to the carcinogenicity of HPV integration.

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Section: Discussionsupporting

confidence: 83%

Long-distance interaction of the integrated HPV fragment with MYC gene and 8q24.22 region upregulating the allele-specific MYC expression in HeLa cells

et al. 2017

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“…Whereas, we specifically target E6 by RNAi, treatment with IP10 resulted in the inhibition of both E6 and E7 expression. Yet, in contrast to studies indicating that combined E6 and E7 inhibition primarily results in senescence (Goodwin et al, 2000;Wells et al, 2000;Hall and Alexander, 2003), induction of apoptosis was reported. In addition, IP10 is likely to exert additional biological activities besides affecting E6/E7 expression.…”

Section: Discussioncontrasting

confidence: 65%

“…E6 targets the PUMA/Bax Pathway M Vogt et al inhibition of E6 and E7. The combined inhibition of E6 and E7 primarily leads to growth arrest and senescence (Goodwin et al, 2000;Wells et al, 2000;Hall and Alexander, 2003), due to the release of pRb from the inhibitory influence of E7 (Psyrri et al, 2004). However, if growth arrest cannot be induced -due to the sustained inactivation of pRb-associated pathways by E7 -inhibition of E6 alone will result in Bax-induced apoptosis.…”

Section: Discussionmentioning

confidence: 99%

Inhibition of Bax activity is crucial for the antiapoptotic function of the human papillomavirus E6 oncoprotein

et al. 2006

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Oncogenic types of human papillomaviruses (HPVs) cause cervical cancer in humans. The antiapoptotic viral E6 gene has been identified as a key factor for maintaining the viability of HPV-positive cancer cells. Although E6 has the potential to modulate many apoptosis regulators, the crucial apoptotic pathway blocked by endogenous E6 in cervical cancer cells remained unknown. Using RNA interference (RNAi), here, we show that targeted inhibition of E6 expression in cervical cancer cells leads to the transcriptional stimulation of the PUMA promoter, in a p53-dependent manner. This is linked to the activation and translocation of Bax to the mitochondrial membrane, cytochrome c release into the cytosol, and activation of caspase-3, in a PUMA-dependent manner. Moreover, inhibition of Bax expression by RNAi efficiently reverts the apoptotic phenotype, which results from inhibition of E6 expression. Thus, interference with the p53/PUMA/ Bax cascade is crucial for the antiapoptotic function of the viral E6 oncogene in HPV-positive cancer cells.

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“…Because of the complex pattern of the polycistronic E6/ E7 mRNAs that are transcribed from chromosomally integrated HPV genomes in cervical cancer cells, it is possible to generate siRNAs that either block E6 expression alone or E6/ E7 expression in combination. 20,35 The selective inhibition of E6 by RNAi is linked to induction of apoptosis. 20 As exosome preparations can be contaminated by material coming from apoptotic cells, 30 which can lead to data misinterpretation, 45 we focused on the combined silencing of viral E6 and E7 expression by RNAi.…”

Section: Discussionmentioning

confidence: 99%

Silencing of human papillomavirus (HPV) E6/E7 oncogene expression affects both the contents and the amounts of extracellular microvesicles released from HPV‐positive cancer cells

Honegger

Leitz

Bulkescher

et al. 2013

Intl Journal of Cancer

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The human papillomavirus (HPV) E6/E7 oncogenes play a crucial role in the HPV-induced carcinogenesis. In this study, the authors investigated whether silencing of endogenous HPV E6/E7 expression may influence the contents or amounts of extracellular microvesicles (eMVs) released from HPV-positive cancer cells. It was found that eMVs secreted from HeLa cells are enriched for Survivin protein. RNA interference studies revealed that maintenance of both intracellular and microvesicular Survivin amounts was strongly dependent on continuous E6/E7 expression. This indicates that intracellular HPV activities are translated into visible alterations of protein contents in eMVs. Besides Survivin, eMVs from HeLa cells contain additional members of the inhibitor of apoptosis protein (IAP) family (XIAP, c-IAP1 and Livin). In contrast, no evidence for the presence of the HPV E6 and E7 oncoproteins in eMVs was obtained. Moreover, it was found that silencing of HPV E6/E7 expression led to a significant increase of exosomes-representing eMVs of endocytic origin-released from HeLa cells. This effect was associated with the reinduction of p53, stimulation of the p53 target genes TSAP6 and CHMP4C that can enhance exosome production and induction of senescence. Taken together, these results show that silencing of HPV E6/E7 oncogene expression profoundly affects both the composition and amounts of eMVs secreted by HPV-positive cancer cells. This indicates that HPVs can induce molecular signatures in eMVs that may affect intercellular communication and could be explored for diagnostic purposes.Specific types of human papillomaviruses (HPVs), such as HPV16 and HPV18, cause cervical cancer and are closely linked to the development of additional human malignancies in the oropharyngeal and anogenital regions. 1 The viral E6 and E7 oncoproteins are crucial both for the HPV-associated induction of transformation and for the maintenance of the tumorigenic phenotype of HPV-positive cervical cancer cells. 2,3 E6 and E7 dysregulate intracellular pathways involved in the control of cellular proliferation, apoptosis and genetic stability. For example, E6 induces the proteolytic degradation of the p53 tumor suppressor protein 4 and stimulates telomerase activity, 5 whereas E7 interferes with the activity of the retinoblastoma tumor suppressor protein, pRb, and other pocket proteins. 6 In contrast to the increasing understanding of the intracellular activities of the viral E6/E7 oncogenes, surprisingly little is known about possible effects on the intercellular communication of HPV-positive cancer cells.Extracellular microvesicles (eMVs) include exosomes that are small vesicles (50-100 nm in diameter) of endosomal origin secreted by a variety of cells, including tumor cells. 7 Exosomes have recently gained much interest in oncology, particularly due to three properties: (i) exosomes secreted from tumor cells can suppress the immune response toward the tumor, 8,9 (ii) tumor cell-derived exosomes can accelerate tumor growth and invasiveness by horizon...

show abstract

RNA Interference of Human Papillomavirus Type 18 E6 and E7 Induces Senescence in HeLa Cells

Cited by 184 publications

References 17 publications

Long-distance interaction of the integrated HPV fragment with MYC gene and 8q24.22 region upregulating the allele-specific MYC expression in HeLa cells

Long-distance interaction of the integrated HPV fragment with MYC gene and 8q24.22 region upregulating the allele-specific MYC expression in HeLa cells

Inhibition of Bax activity is crucial for the antiapoptotic function of the human papillomavirus E6 oncoprotein

Silencing of human papillomavirus (HPV) E6/E7 oncogene expression affects both the contents and the amounts of extracellular microvesicles released from HPV‐positive cancer cells

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