Analysis of genetic control elements in eukaryotes: transcriptional activity or nuclear hitchhiking?
A common way to analyse basal and stimulated activity of eukaryotic genetic control elements, such as promoters and enhancers, is to introduce them into cells via DNA vectors containing an easily assayable reporter gene. Activity is then studied by measurement of transiently produced mRNA or reporter protein. In such assays, it is assumed that the variable measured is proportional to the transcriptional activity of the control element under investigation. Here we question the validity of this generally accepted assumption. Specifically, recent observations indicate that control elements, in addition to modulating transgene transcription, can facilitate the nuclear uptake of their carrier plasmids. This process is mediated by transcription factors or other nuclear proteins harbouring nuclear localisation signals, which bind to the control elements in the cytoplasm and transport the DNA into the nucleus through the protein nuclear import machinery. As the number of mRNA transcripts produced for an epi-chromosomally expressed transgene is directly related to its copy number inside the nucleus, such transport activity may lead to substantial overestimation of the transcriptional potency of the control element(s) studied.
Zohar
M
M
Biotechnology-General, Research Division, Rehovot, Israel.
Mesika
A
A
Reich
Z
Z
D021581:Active Transport, Cell Nucleus; D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism*; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins; D019309:Epstein-Barr Virus Nuclear Antigens / Q000378:metabolism; D005057:Eukaryotic Cells / Q000378:metabolism; D005656:Fungal Proteins / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic
D004268:DNA-Binding Proteins; D019309:Epstein-Barr Virus Nuclear Antigens; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; D016328:NF-kappa B; D009687:Nuclear Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D004247:DNA; C417029:EBV-encoded nuclear antigen 1