Quantitative keratinocyte assay detects two biological activities of human papillomavirus DNA and identifies viral types associated with cervical carcinoma.

Schlegel, Richard; Phelps, William C.; Zhang, Yulian; Barbosa, Miguel

doi:10.1002/j.1460-2075.1988.tb03185.x

Cited by 241 publications

(164 citation statements)

References 36 publications

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“…Cell Culture and Plasmids-Primary HFKs were cultured from neonatal foreskins as described (64) and maintained in keratinocyte growth media (Invitrogen), supplemented with gentamycin (50 g/ml). Primary HFKs P8 (passage 8) were infected with amphotropic LXSN retroviruses expressing AU1-tagged HPV-16 WT E6 (E6A), E6A/ 8S9A10T, E6A/⌬9 -13, or 6bE6A, as described elsewhere (37,65).…”

Section: Methodsmentioning

confidence: 99%

The E6AP Ubiquitin Ligase Is Required for Transactivation of the hTERT Promoter by the Human Papillomavirus E6 Oncoprotein

Yuan

et al. 2005

Journal of Biological Chemistry

103

111

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Most human cancer cells display increased telomerase activity that appears to be critical for continued cell proliferation and tumor formation. The E6 protein of malignancy-associated human papillomaviruses increases cellular telomerase in primary human keratinocytes at least partly via transcriptional activation of the telomerase catalytic subunit, hTERT. In the present study, we investigated whether E6AP, a ubiquitin ligase well known for binding and mediating some of the activities of the E6 oncoprotein, participated in the transactivation of the hTERT promoter. Our results demonstrate that E6 mutants that fail to bind E6AP are also defective for increasing telomerase activity and transactivating the hTERT promoter. More importantly, E6AP knock-out mouse cells and small interfering RNA techniques demonstrated that E6AP was required for hTERT promoter transactivation in both mouse and human cells. Neither E6 nor E6AP bound to the hTERT promoter or activated the promoter in the absence of the partner protein. With all transactivation-competent E6 proteins, induction of the hTERT promoter was dependent upon E box elements in the core promoter. It appears, therefore, that E6-mediated activation of the hTERT promoter requires a complex of E6-E6AP to engage the hTERT promoter and that activation is dependent upon Myc binding sites in the promoter. The recruitment of a cellular ubiquitin ligase to the hTERT promoter during E6-mediated transcriptional activation suggests a role for the local ubiquitination (and potential degradation) of promoter-associated regulatory proteins, including the Myc protein.

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Section: Methodsmentioning

confidence: 99%

The E6AP Ubiquitin Ligase Is Required for Transactivation of the hTERT Promoter by the Human Papillomavirus E6 Oncoprotein

Yuan

et al. 2005

Journal of Biological Chemistry

103

111

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“…Cells were maintained in serum-free keratinocyte growth medium (Life Technologies). Differentiation was induced by allowing for keratinocyte growth in high calcium-containing medium (2.0 mM calcium) consisting of Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum, penicillin, streptomycin, and L-glutamine as described previously (39). Cells were harvested for RNA isolation at different times after induction.…”

Section: Sequence 19mentioning

confidence: 99%

Characterization of ESE-2, a Novel ESE-1-related Ets Transcription Factor That Is Restricted to Glandular Epithelium and Differentiated Keratinocytes

Oettgen¹,

Kas²,

Dube³

et al. 1999

Journal of Biological Chemistry

118

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Epithelial cell differentiation is tightly controlled by distinct sets of transcription factors that regulate the expression of stage-specific genes. We recently isolated the first epithelium-specific Ets transcription factor (ESE-1). Here we describe the characterization of ESE-2, a second epithelium-restricted ESE-1-related Ets factor. Like ESE-1, ESE-2 is induced during keratinocyte differentiation. However, whereas ESE-1 is expressed in the majority of epithelial cell types, ESE-2 expression is restricted to differentiated keratinocytes and glandular epithelium such as salivary gland, prostate, mammary gland, and kidney. In contrast to ESE-1, full-length ESE-2 binds poorly to DNA due to the presence of a negative regulatory domain at the amino terminus. Furthermore, although ESE-1 and the amino-terminally deleted ESE-2 bind with similar affinity to the canonical E74 Ets site, ESE-2 and ESE-1 differ strikingly in their relative affinity toward binding sites in the c-MET and PSMA promoters. Similarly, ESE-1 and ESE-2 drastically differ in their ability to transactivate epitheliumspecific promoters. Thus, ESE-2, but not ESE-1, transactivates the parotid gland-specific PSP promoter and the prostate-specific PSA promoter. In contrast, ESE-1 transactivates the keratinocyte-specific SPRR2A promoter Ets site and the prostate-specific PSMA promoter significantly better than ESE-2. Our results demonstrate the existence of a unique class of related epitheliumspecific Ets factors with distinct functions in epithelial cell gene regulation.Normal epithelial cell development, proliferation, and differentiation are induced by mesenchymal-epithelial interactions and involve cell-cell interactions, extracellular matrix, and soluble growth and differentiation factors. These interactions trigger the activation or expression of a distinct set of transcription factors leading to a specific pattern of gene expression along a tightly controlled pathway. Abnormalities in this process due to deregulated gene expression can lead to the development of benign adenomas or malignant carcinomas that make up the majority of solid tumors. In order to understand tumor development, it is therefore critical to understand normal epithelial cell differentiation and proliferation. Whereas rapid progress in understanding immune system development and gene regulation has led to the discovery and characterization of a whole set of genes involved in leukemia and lymphoma development, relatively little is known about epithelial cell differentiation and organ development and the mechanisms involved in solid tumor formation. Most of the genes involved in chromosomal translocations in leukemias and lymphomas encode transcription factors that under normal physiological conditions coordinate the correct spatial and temporal expression of genes (1). We therefore postulate that similar mechanisms of oncogenesis play a role in epithelium-derived tumors as well. It is thus surprising that many aspects of epithelium-specific gene expression have not been explore...

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“…Oncogenic genital HPV types are capable of immortalizing human primary epithelial cells in culture (DuÈ rst et al, 1987;Pirisi et al, 1987;Tsutsumi et al, 1992;Woodworth et al, 1988). This property is not shared with HPV6 or HPV11 which generally induce benign lesions only (Schlegel et al, 1988;Woodworth et al, 1989). HPVimmortalized keratinocytes can acquire a malignant phenotype after transfection with an activated ras-oncogene or by a combination of gamma-irradiation and long term passage in culture or even long term passage alone (DuÈ rst et al, 1989Chen et al, 1993).…”

Section: Introductionmentioning

confidence: 99%

Identification of novel molecular markers which correlate with HPV-induced tumor progression

et al. 1998

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High risk HPVs (human papillomaviruses) are causally involved in the development of cervical cancer. However, other factors, such as somatic genetic alterations also play a decisive role in malignant conversion of cervical keratinocytes. Mutations and chromosomal aberrations are known to in¯uence the pattern of gene expression. Therefore we used the recently developed RT ± PCR based method of di erential display of mRNAs to detect di erences in gene expression which correlate with tumorigenicity. Non-tumorigenic HPV16-immortalized human foreskin keratinocytes (HPK IA) were compared with their tumorigenic counterparts and 49 di erent genes were identi®ed which were either up-or downregulated. The identi®ed genes encode proteins of the cytoskeleton and the extracellular matrix, the nuclear splicing apparatus, transcription regulators and membrane-associated proteins. The expression pattern of all genes was also examined by RNA ± RNA in situ hybridization in biopsies of normal cervical epithelium, precancerous lesions and cancers. Two genes, C4.8 and C21.7, are of particular interest because their expression is upregulated in a subset of high grade precancerous lesions and in over 58% of cancers. These two genes may therefore be considered as putative progression markers. C4.8 is a new member of the transmembrane 4 (TM-4) protein family which includes tumor-associated antigens such as CD63 and TAPA-1, whereas C21.7 shows no signi®cant homologies to any known genes or proteins.

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Quantitative keratinocyte assay detects two biological activities of human papillomavirus DNA and identifies viral types associated with cervical carcinoma.

Cited by 241 publications

References 36 publications

The E6AP Ubiquitin Ligase Is Required for Transactivation of the hTERT Promoter by the Human Papillomavirus E6 Oncoprotein

The E6AP Ubiquitin Ligase Is Required for Transactivation of the hTERT Promoter by the Human Papillomavirus E6 Oncoprotein

Characterization of ESE-2, a Novel ESE-1-related Ets Transcription Factor That Is Restricted to Glandular Epithelium and Differentiated Keratinocytes

Identification of novel molecular markers which correlate with HPV-induced tumor progression

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