DNA damage sensible engineered promoter for cellular biosensing of cytotoxicity.
We have established a cytotoxic sensor cell line by transfecting HepG2 cells with a luciferase protein plasmid derived from the heat shock protein 70B' (HSP70B') promoter, which is induced by cytotoxic reagents. HSP70B genes are up-regulated by a wide-range of cytotoxic stimulators, in particular, those that denature proteins. However, the HSP70B genes do not respond to DNA damage. We used a PCR array to detect marker genes of DNA damage-related cytotoxic stimulation and found the BTG2 gene to be one such gene. Analysis of the BTG2 gene functional promoter region by transfection of various deletion constructs into HepG2 cells indicated that the p53 and NFY biding sites on BTG2 are important for the response to DNA damage. We then constructed HepG2 sensor cells using the functional BTG2 promoter, and found that these sensor cells can specifically detect the cytotoxicity accompanied by DNA strand breaks with high sensitivity.
Wada
Ken-Ichi
K
Advanced Medical Materials Group, Biomaterials Center, National Institute for Materials Sciences, 1-1 Namiki, Tsukuba, Ibaraki 305-0044, Japan.
Hamaguchi
Yu
Y
Furukawa
Kiyoshi
K
Taniguchi
Akiyoshi
A
D015374:Biosensing Techniques / Q000379:methods*; D002460:Cell Line; D002477:Cells / Q000187:drug effects*; D003603:Cytotoxins / Q000633:toxicity*; D004249:DNA Damage; D017930:Genes, Reporter; D016147:Genes, Tumor Suppressor; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D011401:Promoter Regions, Genetic; D012680:Sensitivity and Specificity; D025521:Tumor Suppressor Proteins
D003603:Cytotoxins; D017874:Immediate-Early Proteins; D025521:Tumor Suppressor Proteins; C070114:BTG2 protein, human; D008156:Luciferases