Different mechanisms contribute to the E2-mediated transcriptional repression of human papillomavirus type 18 viral oncogenes.
Transcription of the human papillomavirus type 18 (HPV18) E6 and E7 oncogenes is repressed by the viral E2 protein. In C33 cells, we have previously shown that of the four E2 binding sites (E2 BS) present in the HPV18 long control region (LCR), only the binding site adjacent to the TATA box (E2 BS 1) was involved in E2-mediated repression. In the present study, we sought to determine whether this phenomenon was conserved in other cell lines. We first showed that all three E2 BS proximal to the P105 promoter were required for full repression of its activity in HeLa and HaCaT cells. Repression by E2 at E2 BS 2 occurred through the displacement of Sp1. Second, a truncated E2 product, lacking the N-terminal transactivation domain, repressed transcription more efficiently than the full-length protein. Repression was abolished when the N-terminal domain of E2 was replaced by the activation domain of VP16. The VP16-E2 chimeric protein could activate transcription from an LCR mutated in its TATA box. DNA-protein binding studies showed that E2 associates with its four binding sites in the LCR with similar affinities. However, challenge of such complexes with excess binding sites demonstrated that interaction with E2 BS 4 was the most stable while interaction with E2 BS 1 was the least stable. Furthermore, complexes with the full-length E2 were less stable than those formed with the N-terminally truncated protein.
Demeret
C
C
Département des Biotechnologies, URA 1644 du CNRS, Institut Pasteur, Paris, France.
Desaintes
C
C
Yaniv
M
M
Thierry
F
F
D001665:Binding Sites; D002460:Cell Line; D004268:DNA-Binding Proteins; D015967:Gene Expression Regulation, Viral; D005814:Genes, Viral; D006367:HeLa Cells; D006801:Humans; D009856:Oncogene Proteins, Viral / Q000235:genetics / Q000378:metabolism*; D009857:Oncogenes; D027383:Papillomaviridae / Q000235:genetics*; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D016329:Sp1 Transcription Factor / Q000378:metabolism; D014158:Transcription, Genetic
D004268:DNA-Binding Proteins; C052603:E6 protein, Human papillomavirus type 18; C052633:E7 protein, Human papillomavirus type 18; D009856:Oncogene Proteins, Viral; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D016329:Sp1 Transcription Factor; C069102:oncogene protein E2, Human papillomavirus type 18