Modulation of Reporter EGFP Gene Expression by a Disease-Associated Human Intra-Intronic Minisatellite Upon Transient and Stable Transfection
Year: 2013
Type: article
Abstract: Currently, our understanding of associations between minisatellite poly morphisms and diseases is far fro m clear. This is in part due to great differences in human and animal min isatellite structure. Thus, here we have used plasmids which contained the reporter gene EGFP under eukaryotic pro moter ROSA 26, and different allelles of hu man minisatellite UPS29 for both transient and stable transfection of F9 cells induced to differentiate, but not terminally d ifferentiated, to establish whether the min isatellite would be capable of affecting exp ression of EGFP in the episomal state and in a closer to native chromatin environment. Upon transient transfection, we found enhanced reporter gene expression for constructs with minisatellite alleles inserts. By contrast, in cells that were stably transfected UPS29 alleles suppressed reporter construct expression compared to controls with no inserted UPS29 allele. The most pro minent effect was observed for the shortest UPS29 allele associated with diseases. For stably transfected cell lines, integrated transgene copy number and some epigenetic characteristics of nuclei in situ did not differ among constructs containing different UPS29 alleles. The enhancer effect seen with transient transfection is speculated to be due to accessibility to specific transcription factors. The suppressive effect seen in stable transfectants might be determined mainly by methylation of minisatellite sequences rather than by peculiarities of host genome.
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: diamond
OpenAlex ID: W2096924256