Abstract
BackgroundTetracycline regulated ectopic gene expression is a widely used tool to study gene function. However, the tetracycline regulator (tetR) itself has been reported to cause certain phenotypic changes in mammalian cells. We, therefore, asked whether human myeloid U937 cells expressing the tetR in an autoregulated manner would exhibit alterations in gene expression upon removal of tetracycline.Methodology/Principal FindingsMicroarray analyses revealed that 172 and 774 unique genes were significantly differentially expressed by at least 2- or 1.5-fold, respectively, when tetR expressing U937 cells were maintained in media with or without the antibiotic.Conclusions/SignificanceThese alterations in gene expression are likely to contribute to the phenotypic consequences of tetR expression. In addition, they need to be taken into consideration when using the tetR system for the identification of target genes of transcription factors or other genes of interest.
Highlights
Inducible ectopic expression is a widely used tool to study gene function
Because we and others have found that tetracycline regulator (tetR) expressing cells show phenotypic changes in response to altered concentrations of antibiotic even if they do not contain a tetracycline regulated gene of interest [5,6,10,12,15], we asked whether these changes may correspond to altered gene expression patterns
Using microarray analysis we found that the levels of several hundred mRNAs were altered significantly after transfer of U937T_pUHD10S cells to tetracycline free media
Summary
Inducible ectopic expression is a widely used tool to study gene function. The expression of genes cloned downstream of a promoter containing several copies of the bacterial tetracycline operator (tetO) can be regulated by removal or addition of tetracycline or tetracycline derivatives from/to the culture media. In further improvements of the system, tetR was placed under the control of tetO elements to create an autoregulatory loop, avoiding problems caused by toxicity of constitutive tetR expression and reducing background levels of the gene of interest in the repressed state [3,4]. Tetracycline regulated ectopic gene expression is a widely used tool to study gene function. We asked whether human myeloid U937 cells expressing the tetR in an autoregulated manner would exhibit alterations in gene expression upon removal of tetracycline
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