The p65 domain from NF-kappaB is an efficient human activator in the tetracycline-regulatable gene expression system.
The tc-responsive TetR protein allows the investigation of various transcriptional activators in respective fusion proteins. We have fused eight well-known human activator domains to the C-terminus of TetR and determined the properties of the resulting transactivators using a tetracycline-responsive promoter in three human cell lines (HeLa, BJAB, and Jurkat). Several-hundred-fold activation was exclusively obtained with the acidic p65 domain from NF-kappaB and with VP16, which served as a positive control. In contrast, at least 10-fold lower factors of activation were achieved with ITF-1, ITF-2, and MTF-1. The induction properties of the p65 domain are identical to those of VP16 in all three human cell lines and when fused to the reverse TetR. The combination of the novel reverse p65 fusion with the TetR(B/E)-KRAB construct resulted in active silencing and full activation. This is the first report of an expression system with minimal basal activity and high induction levels without viral protein domains.
Urlinger
S
S
Lehrstuhl für Mikrobiologie, Friedrich-Alexander-Universität Erlangen-Nürnberg, Staudtstr. 5, D-91058, Erlangen, Germany.
Helbl
V
V
Guthmann
J
J
Pook
E
E
Grimm
S
S
Hillen
W
W
D000818:Animals; D001402:B-Lymphocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D001665:Binding Sites / Q000235:genetics; D005786:Gene Expression Regulation / Q000187:drug effects; D006367:HeLa Cells; D017405:Herpes Simplex Virus Protein Vmw65 / Q000235:genetics / Q000378:metabolism; D006801:Humans; D019169:Jurkat Cells; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D016328:NF-kappa B / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D013601:T-Lymphocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D013752:Tetracycline / Q000494:pharmacology*; D013753:Tetracycline Resistance / Q000235:genetics; D015534:Trans-Activators; D051996:Transcription Factor RelA; D014158:Transcription, Genetic / Q000187:drug effects; D014407:Tumor Cells, Cultured
D017405:Herpes Simplex Virus Protein Vmw65; D016328:NF-kappa B; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; D051996:Transcription Factor RelA; D008156:Luciferases; D013752:Tetracycline