Abstract:Our results unraveled the possibility that HPV-16 E6/E7 could promote cell invasive potential via regulating cadherin switching, and consequently contribute to progression and metastasis of cervical cancer.
“…4a). These pathways are consistent with known biological activities of E7, which has been reported to regulate multiple genes to modulate translation 48 , promote cell migration 49 , play a role in macromolecular complex subunit organization 50 , DNA replication 51 and redox balancing 52 .Figure 4Label-free mass spectrometry quantification of RPE1 vector and E7 expressing cells under hypoxia. ( a ) RPE1 vector and E7 expressing cells treated with 200 μM DFO were prepared for mass spectrometry.…”
The human papillomavirus (HPV) plays a central role in cervical carcinogenesis and its oncogene E7 is essential in this process. We showed here that E7 abrogated the G1 cell cycle checkpoint under hypoxia and analyzed key cell cycle related proteins for their potential role in this process. To further explore the mechanism by which E7 bypasses hypoxia-induced G1 arrest, we applied a proteomic approach and used mass spectrometry to search for proteins that are differentially expressed in E7 expressing cells under hypoxia. Among differentially expressed proteins identified, Cdc6 is a DNA replication initiation factor and exhibits oncogenic activities when overexpressed. We have recently demonstrated that Cdc6 was required for E7-induced re-replication. Significantly, here we showed that Cdc6 played a role in E7-mediated G1 checkpoint abrogation under hypoxic condition, and the function could possibly be independent from its role in DNA replication initiation. This study uncovered a new function of Cdc6 in regulating cell cycle progression and has important implications in HPV-associated cancers.
“…4a). These pathways are consistent with known biological activities of E7, which has been reported to regulate multiple genes to modulate translation 48 , promote cell migration 49 , play a role in macromolecular complex subunit organization 50 , DNA replication 51 and redox balancing 52 .Figure 4Label-free mass spectrometry quantification of RPE1 vector and E7 expressing cells under hypoxia. ( a ) RPE1 vector and E7 expressing cells treated with 200 μM DFO were prepared for mass spectrometry.…”
The human papillomavirus (HPV) plays a central role in cervical carcinogenesis and its oncogene E7 is essential in this process. We showed here that E7 abrogated the G1 cell cycle checkpoint under hypoxia and analyzed key cell cycle related proteins for their potential role in this process. To further explore the mechanism by which E7 bypasses hypoxia-induced G1 arrest, we applied a proteomic approach and used mass spectrometry to search for proteins that are differentially expressed in E7 expressing cells under hypoxia. Among differentially expressed proteins identified, Cdc6 is a DNA replication initiation factor and exhibits oncogenic activities when overexpressed. We have recently demonstrated that Cdc6 was required for E7-induced re-replication. Significantly, here we showed that Cdc6 played a role in E7-mediated G1 checkpoint abrogation under hypoxic condition, and the function could possibly be independent from its role in DNA replication initiation. This study uncovered a new function of Cdc6 in regulating cell cycle progression and has important implications in HPV-associated cancers.
“…More importantly, cadherin switches (referred to as switches from E-cadherin to P-cadherin or E-cadherin to N-cadherin) have been regarded as key hallmarks of EMT [28] and to be involved in TGF-β signaling-induced EMT [29]. Previously, we found that aberrant E-cadherin expression led to increased migration and invasion of cervical SCC cells and could serve as a key biomarker for a poor prognosis of early-stage cervical SCC [7,8]. Furthermore, we found that P-cadherin could affect the expression of E-cadherin and p120 catenin, thereby contributing to cervical cancer initiation and progression [30].…”
Section: Discussionmentioning
confidence: 97%
“…Previously, we found that HPV-16 E6/E7 may regulate an E-P cadherin switch to promote cervical SCC cell growth and invasion [8].…”
Section: Hpv-16 E6/e7 Simultaneously Regulates Mir-34a and Wnt1 Exprementioning
confidence: 97%
“…In our previous study, aberrantly expressed classical cadherins, including epithelial cadherin (E-cadherin) and placental cadherin (P-cadherin), were detected in cervical lesions and to be significantly associated with a poor prognosis and survival in early-stage cervical squamous cell carcinoma (SCC) patients [7]. In addition, we found that exogenous expression of HPV-16 E6/E7 in cervical cell lines significantly increased P-cadherin levels and decreased Ecadherin levels, consequently leading to an E-cadherin to Pcadherin switch (E-P cadherin switch) [8]. These results indicate that HPV-16 E6/E7 may be involved in cervical cancer initiation and progression by regulating the E-P cadherin switch.…”
Purpose Persistent infection with high-risk human papillomavirus (HR-HPV) is thought to play a prominent role in the initiation and progression of almost all cases of cervical cancer. Previously, we and others found that microRNA 34a (miR-34a) may be regulated by HR-HPV E6 to contribute to the development of cervical cancer. Here, we aimed to identify the oncogenic potential and clinical significance of a known miR-34a target, WNT1, in cervical squamous cell carcinoma (SCC) development and to investigate the associated mechanisms underlying cervical SCC cell proliferation and invasion. Methods WNT1 and miR-34a expression levels were assessed in primary cervical lesions using immunohistochemistry and qRT-PCR, respectively. The cellular effects and the expression of its associated genes were examined in cervical SCC-derived Siha and Caski cells after siRNA-WNT1 (downregulation) or miR-34a mimic (upregulation) treatment. A cervical SCC xenograft mouse model was used to investigate the in vivo effects of miR-34a overexpression. HPV-16 E6/E7 expression was inhibited by gene promoter siRNA targeting, after which the levels of miR-34a and WNT1 were examined. Results WNT1 protein upregulation was found to be associated with a poor prognosis in cervical SCC patients. In vitro assays in Siha and Caski cells revealed that WNT1 downregulation decreased cell proliferation and invasion, inhibited WNT/β-catenin activation and affected the expression of E-cadherin and P-cadherin. MiR-34a upregulation resulted in decreased WNT1 expression. An inverse correlation between miR-34a and WNT1 expression was also observed in primary cervical SCC tissues. In addition, we found that MiR-34a could regulate an E-cadherin to P-cadherin switch (E-P cadherin switch) to inhibit cell proliferation and tumorigenesis in vitro and in vivo via inactivation of the WNT1/β-catenin pathway. Finally, we found that decreased HPV-16 E6/E7 expression resulted in miR-34a upregulation and WNT1 downregulation in Siha and Caski cells. Conclusions From our results we conclude that WNT1, as a target of miR-34a, can promote cervical SCC cell proliferation and invasion by induction of an E-P cadherin switch via the WNT1/β-catenin pathway. Our results may provide new options for the treatment of patients with cervical SCC.Keywords Cervical squamous cell carcinoma . miR-34a . WNT1 . E-cadherin . P-cadherin . Cadherin switch Electronic supplementary material The online version of this article (https://doi.
“…Although similar permeable membrane insert systems have been employed for the study of secreted factors involved in viral infections, cancer biology, and immune cell migration [22][23][24][25][26] , few studies involving bacterial interactions with host cells have utilized this approach 6,27,28 . Even studies that have employed such systems to explore interactions between bacteria and host cells have primarily focused on migration of inflammatory cells or bacteria through an epithelial or endothelial monolayer plated on the permeable membrane insert.…”
Many bacterial pathogens secrete potent toxins to aid in the destruction of host tissue, to initiate signaling changes in host cells or to manipulate immune system responses during the course of infection. Though methods have been developed to successfully purify and produce many of these important virulence factors, there are still many bacterial toxins whose unique structure or extensive post-translational modifications make them difficult to purify and study in in vitro systems. Furthermore, even when pure toxin can be obtained, there are many challenges associated with studying the specific effects of a toxin under relevant physiological conditions. Most in vitro cell culture models designed to assess the effects of secreted bacterial toxins on host cells involve incubating host cells with a one-time dose of toxin. Such methods poorly approximate what host cells actually experience during an infection, where toxin is continually produced by bacterial cells and allowed to accumulate gradually during the course of infection. This protocol describes the design of a permeable membrane insert-based bacterial infection system to study the effects of Streptolysin S, a potent toxin produced by Group A Streptococcus, on human epithelial keratinocytes. This system more closely mimics the natural physiological environment during an infection than methods where pure toxin or bacterial supernatants are directly applied to host cells. Importantly, this method also eliminates the bias of host responses that are due to direct contact between the bacteria and host cells. This system has been utilized to effectively assess the effects of Streptolysin S (SLS) on host membrane integrity, cellular viability, and cellular signaling responses. This technique can be readily applied to the study of other secreted virulence factors on a variety of mammalian host cell types to investigate the specific role of a secreted bacterial factor during the course of infection.
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