Transcriptional activity in 3T3, F9, and PCC4 embryonal carcinoma cells: a systematic deletion and linker-scanning study of the polyomavirus enhancer.
The polyomavirus (PyV) genome is not expressed in undifferentiated embryonal carcinoma (EC) cells such as PCC4 or F9 EC cells. All the viral mutants that have been selected for their expression in these cells harbor mutations or rearrangements within a region which is important for early and late transcription (as transcriptional enhancer) as well as for viral DNA replication. We have studied the role of the different parts of this enhancer on the transcription driven by early PyV promoter. Two series of constructions containing progressive deletions starting from the ends of the region and one series containing linker-scanning mutations were tested in a luciferase assay in three cell lines: 3T3, F9, or PCC4. The results revealed that some regions that have not yet been shown to bind transcription factors are nevertheless important for transcriptional activity. Two subregions between nucleotides 5179 and 5187 and between 5220 and 5227 were found to be inhibitory for the activity of the enhancer in EC cells. The PEA1/AP1 binding site was also unexpectedly shown to be important in F9 EC cells.
Laboratoire Virus et Différenciation, Université Pierre et Marie Curie, UPR 272, Centre National de la Recherche Scientifique, Villejuif, France.
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D016475:3T3 Cells; D000818:Animals; D029681:Arabidopsis Proteins; D001483:Base Sequence; D001665:Binding Sites; D018236:Carcinoma, Embryonal; D002460:Cell Line; D004742:Enhancer Elements, Genetic; D015967:Gene Expression Regulation, Viral; D066298:In Vitro Techniques; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010940:Plant Proteins / Q000378:metabolism; D011120:Polyomavirus / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D017384:Sequence Deletion; D013329:Structure-Activity Relationship; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic
D029681:Arabidopsis Proteins; C084017:PEA1 protein, Arabidopsis; D010940:Plant Proteins; D012333:RNA, Messenger; D014157:Transcription Factors; C065788:transcription factor PEA3