A chromosomal position effect on gene targeting in human cells.
We describe gene targeting experiments involving a human cell line (RAN10) containing, in addition to its endogenous alleles, two ectopic alleles of the interferon-inducible gene 6-16. The frequency of gene targeting at one of the ectopic 6-16 alleles (H3.7) was 34-fold greater than the combined frequency of gene targeting involving endogenous 6-16 alleles in RAN10. Preference for H3.7 was maintained when the target loci in RAN10 were transcriptionally activated by interferon. Despite the 34-fold preference for H3.7, the absolute gene targeting efficiency in RAN10 was only 3-fold higher than in the parental HT1080 cell line. These data suggest that different alleles can compete with each other, and perhaps with non-homologous loci, in a step which is necessary, but not normally rate-limiting, for gene targeting. The efficiency of this step can therefore be more sensitive to chromosomal position effects than the rate-determining steps for gene targeting. The nature of the position effects involved remains unknown but does not correlate with transcription status, which in our system has a very modest influence on the frequency of gene targeting. In summary, our work unequivocally identifies a position effect on gene targeting in human cells.
Yáñez
Rafael J
RJ
Gene Targeting Group, MRC Clinical Sciences Centre, Faculty of Medicine, Imperial College, Hammersmith Hospital Campus, Du Cane Road, London W12 0NN, UK.
Porter
Andrew C G
AC
D000483:Alleles; D002877:Chromosomes, Human; D002999:Clone Cells; D004791:Enzyme Inhibitors / Q000494:pharmacology; D018390:Gene Targeting / Q000379:methods*; D056572:Histone Deacetylase Inhibitors; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D007372:Interferons / Q000494:pharmacology; D024101:Mitochondrial Proteins; D011123:Polyploidy; D011506:Proteins / Q000235:genetics; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured; D015854:Up-Regulation
D004791:Enzyme Inhibitors; D056572:Histone Deacetylase Inhibitors; D006877:Hydroxamic Acids; C468762:IFI6 protein, human; D024101:Mitochondrial Proteins; D011506:Proteins; C012589:trichostatin A; D007372:Interferons