Reversal of the silencing of tetracycline-controlled genes requires the coordinate action of distinctly acting transcription factors.

Details

Serval ID
serval:BIB_0CF2945FAA97
Type
Article: article from journal or magazin.
Collection
Publications
Title
Reversal of the silencing of tetracycline-controlled genes requires the coordinate action of distinctly acting transcription factors.
Journal
Journal of Gene Medicine
Author(s)
Pankiewicz R., Karlen Y., Imhof M.O., Mermod N.
ISSN
1099-498X[print], 1099-498X[linking]
Publication state
Published
Issued date
01/2005
Volume
7
Number
1
Pages
117-132
Language
english
Notes
Publication types: Journal Article ; Research Support, Non-U.S. Gov't
Publication Status: ppublish
Abstract
BACKGROUND: Regulation of genes transferred to eukaryotic organisms is often limited by the lack of consistent expression levels in all transduced cells, which may result in part from epigenetic gene silencing effects. This reduces the efficacy of ligand-controlled gene switches designed for somatic gene transfers such as gene therapy. METHODS: A doxycycline-controlled transgene was stably introduced in human cells, and clones were screened for epigenetic silencing of the transgene. Various regulatory proteins were targeted to the silent transgene, to identify those that would mediate regulation by doxycycline. RESULTS: A doxycycline-controlled minimal promoter was found to be prone to gene silencing, which prevents activation by a fusion of the bacterial TetR DNA-binding domain with the VP16 activator. DNA modification studies indicated that the silenced transgene adopts a poorly accessible chromatin structure. Several cellular transcriptional activators were found to restore an accessible DNA structure when targeted to the silent transgene, and they cooperated with Tet-VP16 to mediate regulation by doxycycline. CONCLUSIONS: Reversal of the silencing of a tetracycline-regulated minimal promoter requires a chromatin-remodeling activity for subsequent promoter activation by the Tet-VP16 fusion protein. Thus, distinct regulatory elements may be combined to obtain long-term regulation and persistent expression of exogenous genes in eukaryotic cells.
Keywords
Cell Line, Chromatin/metabolism, Clone Cells, DNA Methylation, Epigenesis, Genetic/drug effects, Gene Expression Regulation/drug effects, Gene Silencing, Gene Therapy, Genes, Reporter, Humans, Luciferases/metabolism, Promoter Regions, Genetic, Recombinant Fusion Proteins/metabolism, Tetracycline/pharmacology, Trans-Activators/genetics, Transcription Factors/genetics, Transcription Factors/metabolism, Transfection, Transgenes, beta-Galactosidase/analysis, beta-Galactosidase/metabolism
Pubmed
Web of science
Create date
24/01/2008 10:41
Last modification date
20/08/2019 12:34
Usage data