Down-regulation of transforming growth factor-β type II receptor (TGF-βRII) protein and mRNA expression in cervical cancer

<p>Abstract</p> <p>Background</p> <p>Cervical carcinogenesis is a multistep process initiated by "high risk" human papillomaviruses (HR-HPV), most commonly HPV16. The infection <it>per se </it>is, however, not sufficient to induce malignant conversion. Transforming Growth Factor β (TGF-β) inhibits epithelial proliferation and altered expression of TGF-β or its receptors may be important in carcinogenesis. One cofactor candidate to initiate neoplasia in cervical cancer is the prolonged exposure to sex hormones. Interestingly, previous studies demonstrated that estrogens suppress TGF-β induced gene expression. To examine the expression of TGF-β2, TGF-βRII, p15 and c-myc we used <it>in situ </it>RT-PCR, real-time PCR and immunohistochemistry in transgenic mice expressing the oncogene E7 of HPV16 under control of the human Keratin-14 promoter (K14-E7 transgenic mice) and nontransgenic control mice treated for 6 months with slow release pellets of 17β-estradiol.</p> <p>Results</p> <p>Estrogen-induced carcinogenesis was accompanied by an increase in the incidence and distribution of proliferating cells solely within the cervical and vaginal squamous epithelium of K14-E7 mice. TGF-β2 mRNA and protein levels increased in K14-E7 transgenic mice as compared with nontransgenic mice and further increased after hormone-treatment in both nontransgenic and transgenic mice. In contrast, TGF-βRII mRNA and protein levels were decreased in K14-E7 transgenic mice compared to nontransgenic mice and these levels were further decreased after hormone treatment in transgenic mice. We also observed that c-myc mRNA levels were high in K14-E7 mice irrespective of estrogen treatment and were increased in estrogen-treated nontransgenic mice. Finally we found that p15 mRNA levels were not increased in K14-E7 mice.</p> <p>Conclusion</p> <p>These results suggest that the synergy between estrogen and E7 in inducing cervical cancer may in part reflect the ability of both factors to modulate TGF-β signal transduction.</p

Heparin (GAG-hed) inhibits LCR activity of Human Papillomavirus type 18 by decreasing AP1 binding

BACKGROUND: High risk HPVs are causative agents of anogenital cancers. Viral E6 and E7 genes are continuously expressed and are largely responsible for the oncogenic activity of these viruses. Transcription of the E6 and E7 genes is controlled by the viral Long Control Region (LCR), plus several cellular transcription factors including AP1 and the viral protein E2. Within the LCR, the binding and activity of the transcription factor AP1 represents a key regulatory event in maintaining E6/E7 gene expression and uncontrolled cell proliferation. Glycosaminoglycans (GAGs), such as heparin, can inhibit tumour growth; they have also shown antiviral effects and inhibition of AP1 transcriptional activity. The purpose of this study was to test the heparinoid GAG-hed, as a possible antiviral and antitumoral agent in an HPV18 positive HeLa cell line. METHODS: Using in vivo and in vitro approaches we tested GAG-hed effects on HeLa tumour cell growth, cell proliferation and on the expression of HPV18 E6/E7 oncogenes. GAG-hed effects on AP1 binding to HPV18-LCR-DNA were tested by EMSA. RESULTS: We were able to record the antitumoral effect of GAG-hed in vivo by using as a model tumours induced by injection of HeLa cells into athymic female mice. The antiviral effect of GAG-hed resulted in the inhibition of LCR activity and, consequently, the inhibition of E6 and E7 transcription. A specific diminishing of cell proliferation rates was observed in HeLa but not in HPV-free colorectal adenocarcinoma cells. Treated HeLa cells did not undergo apoptosis but the percentage of cells in G(2)/M phase of the cell cycle was increased. We also detected that GAG-hed prevents the binding of the transcription factor AP1 to the LCR. CONCLUSION: Direct interaction of GAG-hed with the components of the AP1 complex and subsequent interference with its ability to correctly bind specific sites within the viral LCR may contribute to the inhibition of E6/E7 transcription and cell proliferation. Our data suggest that GAG-hed could have antitumoral and antiviral activity mainly by inhibiting AP1 binding to the HPV18-LCR

The E6 Oncoprotein from HPV16 Enhances the Canonical Wnt/?-Catenin Pathway in Skin Epidermis In Vivo

The contribution of the Wnt signaling pathway to human papilloma virus (HPV)-induced carcinogenesis is poorly understood. In high-grade dysplastic lesions that are caused by high-risk HPVs (HR-HPV), ?-catenin is often located in the cell nucleus, which suggests that Wnt pathway may be involved in the development of HPV-related carcinomas. Most of the oncogenic potential of HR-HPVs resides on the PDZ-binding domain of E6 protein. We hypothesized that the PDZ-binding domain of the HPV16-E6 oncoprotein induces the nuclear accumulation of ?-catenin due to its capacity to degrade PDZ-containing cellular targets. To test this hypothesis, we evaluated the staining pattern of ?-catenin in the skin epidermis of transgenic mice expressing the full-length E6 oncoprotein (K14E6 mice) and measured LacZ gene expression in K14E6 mice that were crossed with a strain expressing LacZ that was knocked into the Axin2 locus (Axin2+/LacZ mice). Here, we show that the E6 oncoprotein enhances the nuclear accumulation of ?-catenin, the accumulation of cellular ?-catenin–responsive genes, and the expression of LacZ. None of these effects were observed when a truncated E6 oncoprotein that lacks the PDZ-binding domain was expressed alone (K14E6?PDZ mice) or in combination with Axin2+/LacZ. Conversely, cotransfection with either E6 or E6?PDZ similarly enhanced canonical Wnt signaling in short-term in vitro assays that used a luciferase Wnt/?-catenin/TCF-dependent promoter. We propose that the activation of canonical Wnt signaling could be induced by the HPV16-E6 oncoprotein; however, the participation of the E6 PDZ-binding domain seems to be important in in vivo models only. Mol Cancer Res; 10(2); 250–8. ©2011 AACR

Effect of Sarkosyl on Chromatin and Viral RNA Synthesis: The Isolation of SV40 Transcription Complex

The endogenous RNA polymerase activity of mouse nuclei is enhanced several‐fold by the anionic detergent Sarkosyl. The action of Sarkosyl is exerted primarily on the α‐amanitin sensitive form of the enzyme. This detergent causes the release of nearly all the protein associated with cellular DNA but does not release initiated RNA polymerase. Sarkosyl was also able to activate the RNA polymerase activity from mitotic cells, in which transcription of the highly condensed chromatin is minimal. The use of this anionic detergent has also permited the extraction of a nucleoprotein complex from Simian Virus 40 (SV40) infected monkey cells. Molecular hybridization experiments have established the viral specificity of the RNA synthesized in vitro by the endogenous polymerase present in this complex. Copyright © 1976, Wiley Blackwell. All rights reservedSCOPUS: ar.jinfo:eu-repo/semantics/publishe

Biomedicina molecular, oncogenes y cáncer humano

This article offers a general introduction to the role played by molecular biology as applied to medical research, in our understanding of cancer. In particular, I review recent work carried out by my research group, as well as by other groups around the world, bearing on the relationship between the alterations in certain cellular genes and the phenomenon of cancer, and the role played by certain viruses in the development of human cancers.

Biomedicina molecular, oncogenes y cáncer humano

Se presenta una introducción general a la importancia que tiene la biología molecular aplicada a la investigación médica. En particular se presenta una revisión del trabajo reciente llevado acabo por un grupo de investigación, conducente a establecer la relación entre las alteraciones de ciertos genes celulares y el fenómeno del cáncer; asi como el papel que tienen algunos virus en el desarrollo del cáncer en humanos

CARACTERISATION D'UN COMPLEXE DE TRANSCRIPTION VIRAL

SCOPUS: NotDefined.jinfo:eu-repo/semantics/publishe

Characterization of a soluble simian virus 40 transcription complex

The authors have previously described the isolation, from nuclei of monkey cells infected with Simian virus 40 (SV40), of a nucleoprotein complex which is able to achieve viral transcription. This complex contains SV40 DNA and RNA polymerase II molecules which have initiated transcription during the viral development. The authors show here, by molecular hybridization experiments, that most of the templates active in SV40 transcription can be dissociated from host DNA. In conditions where supercoiled SV40 DNA form I sediments at 21 S, the transcription complex has a sedimentation coefficient of about 25 S. Inhibition of viral DNA synthesis by cytosine arabinonucleoside or chloroquine does not affect the activity of the transcription complex, which suggests that replicating molecules are not required for viral RNA synthesis and that SV40 DNA form I could serve as template for late SV40 transcription. A large fraction of the RNA synthesized in vitro remains associated with the SV40 DNA template in cesium sulfate density gradient. The RNA chains produced by the complex are heterogeneous in size, most of them being as large or larger than the viral genome.SCOPUS: ar.jinfo:eu-repo/semantics/publishe

Isolement et caractérisation d'un minichromosome actif pour la transcription

SCOPUS: NotDefined.jinfo:eu-repo/semantics/publishe