title: The E6AP ubiquitin ligase is required for transactivation of the hTERT promoter by the human papillomavirus E6 oncoprotein.
abstract: 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.
authors.affiliation: Department of Pathology and Oncology, Georgetown University Medical School, 3900 Reservoir Rd. NW, Washington, DC 20057, USA.
authors.forename: Xuefeng
authors.initials: X
authors.lastname: Liu
authors.forename: Hang
authors.initials: H
authors.lastname: Yuan
authors.forename: Baojin
authors.initials: B
authors.lastname: Fu
authors.forename: Gary L
authors.initials: GL
authors.lastname: Disbrow
authors.forename: Tania
authors.initials: T
authors.lastname: Apolinario
authors.forename: Vjekoslav
authors.initials: V
authors.lastname: Tomaic
authors.forename: Melissa L
authors.initials: ML
authors.lastname: Kelley
authors.forename: Carl C
authors.initials: CC
authors.lastname: Baker
authors.forename: Jon
authors.initials: J
authors.lastname: Huibregtse
authors.forename: Richard
authors.initials: R
authors.lastname: Schlegel
mesh_terms: D000111:Acetylcysteine / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D015153:Blotting, Western; D020134:Catalytic Domain; D002460:Cell Line; D002478:Cells, Cultured; D002843:Chromatin / Q000378:metabolism; D047369:Chromatin Immunoprecipitation; D004268:DNA-Binding Proteins; D005347:Fibroblasts / Q000378:metabolism; D005822:Genetic Vectors / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D047468:Immunoprecipitation; D015603:Keratinocytes / Q000378:metabolism; D008156:Luciferases / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D009154:Mutation; D009856:Oncogene Proteins, Viral / Q000378:metabolism*; D027383:Papillomaviridae / Q000235:genetics*; D010957:Plasmids / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D016271:Proto-Oncogene Proteins c-myc / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D012097:Repressor Proteins / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019098:Telomerase / Q000378:metabolism*; D013997:Time Factors; D015533:Transcriptional Activation; D014162:Transfection; D025801:Ubiquitin / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry* / Q000502:physiology*
chemical_list: D002843:Chromatin; D004268:DNA-Binding Proteins; C068651:E6 protein, Human papillomavirus type 16; D009856:Oncogene Proteins, Viral; D016271:Proto-Oncogene Proteins c-myc; D034741:RNA, Small Interfering; D012097:Repressor Proteins; D025801:Ubiquitin; D008156:Luciferases; C471639:UBE3A protein, human; D044767:Ubiquitin-Protein Ligases; D019098:Telomerase; D000111:Acetylcysteine