A designed curved DNA segment that is a remarkable activator of eukaryotic transcription.
To identify artificial DNA segments that can stably express transgenes in the genome of host cells, we built a series of curved DNA segments that mimic a left-handed superhelical structure. Curved DNA segments of 288 bp (T32) and 180 bp (T20) were able to activate transcription from the herpes simplex virus thymidine kinase (tk) promoter by approximately 150-fold and 70-fold, respectively, compared to a control in a transient transfection assay in COS-7 cells. The T20 segment was also able to activate transcription from the human adenovirus type 2 E1A promoter with an 18-fold increase in the same assay system, and also activated transcription from the tk promoter on episomes in COS-7 cells. We also established five HeLa cell lines with genomes containing T20 upstream of the transgene promoter and control cell lines with T20 deleted from the transgene locus. Interestingly, T20 was found to activate transcription in all the stable transformants, irrespective of the locus. This suggests that the T20 segment may allow stable expression of transgenes, which is of importance in many fields, and may also be useful for the construction of nonviral vectors for gene therapy.
Sumida
Noriyuki
N
Department of Biology, Faculty of Science and Engineering, Konan University, Kobe, Japan.
Nishikawa
Jun-ichi
J
Kishi
Haruka
H
Amano
Miho
M
Furuya
Takayo
T
Sonobe
Haruyuki
H
Ohyama
Takashi
T
D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D004247:DNA / Q000737:chemistry / Q000235:genetics*; D005057:Eukaryotic Cells / Q000378:metabolism*; D005822:Genetic Vectors; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic; D014158:Transcription, Genetic; D019076:Transgenes / Q000235:genetics*; D015854:Up-Regulation / Q000235:genetics*
D004247:DNA