Random isolation of gene activator elements from the human genome.
Long-range-acting gene activator elements were randomly isolated from the human genome by functional selection. HeLa cells were transfected with an enhancer trap, a plasmid containing an enhancerless xanthine-guanosine phosphoribosyltransferase (gpt) gene transcribed from the simian virus 40 early promoter, and stably transformed GPT+ cells were selected. From several transformants, human DNA sequences flanking the enhancer trap were cloned. Two gene activators (GA1 and GA2) were found in the cloned human DNAs. GA1 and GA2 showed strong enhancer activity both in a stable transformation assay and in a transient expression assay. They had functional properties similar to those of other known enhancers: GA1 and GA2 activated the expression of a linked gene over distances of at least 5 kilobases both upstream and downstream in an orientation-independent fashion. GA1 may be required for the initial establishment of gene activation but was not essential for the maintenance of active expression. GA1 and GA2 were active not only in HeLa cells but also in other types of human cells, such as neuroblastoma cells. This indicates a limited but relatively broad cell type specificity. The HeLa genome contains multiple copies of GA1, while GA2 exists once in the genome.
D003001:Cloning, Molecular; D004262:DNA Restriction Enzymes; D004742:Enhancer Elements, Genetic; D005796:Genes; D005809:Genes, Regulator; D005814:Genes, Viral; D006367:HeLa Cells / Q000378:metabolism; D006801:Humans; D010430:Pentosyltransferases / Q000235:genetics*; D010957:Plasmids; D011401:Promoter Regions, Genetic; D013539:Simian virus 40 / Q000235:genetics; D014162:Transfection
D010430:Pentosyltransferases; C024827:xanthine phosphoribosyltransferase; D004262:DNA Restriction Enzymes