Construction of X‐ray‐inducible promoters through cis‐acting element elongation and error‐prone polymerase chain reaction

Abstract

A sensitively responsive promoter to radiation could be constructed using this method, possibly leading to the construction of a promoter of interest that could be applied for clinical use.

Regulation of gene expression in prostate cancer cells with an artificially constructed promoter responsive to radiation

“…In this report, we showed that a promoter responsive to radiation in prostate cancer cells was successfully constructed by a method described in our previous study, 18 although a set of cis-acting elements used were somewhat different. Combining a DNA fragment A radiation-responsive artificial promoter A Morii et al constructed by using randomly connected five different cis-acting elements to another DNA fragment containing the TATA-box, we constructed a promoter library.…”

“…In addition, a constructed promoter library tends to contain many promoters responsive to radiation stimulation and thus, it would not be a very difficult task to obtain a useful promoter out of such a library by screening (Figure 1c). 18 Therefore, it should not be impractical to construct promoters for individual cancer treatments. In addition, one promoter could be used for the treatment of multiple types of cancer as it was shown that clone 880-8 was activated by radiation in five cell lines out of 11 different cell lines tested (Figure 2e).…”

“…However, the promoter showing the highest reactivity to radiation was not or only weakly responsive in prostate cancer cell lines. 18 As the promoter was chosen out of a promoter library consisting of only 11 different clones, to expand our possibilities of generating a promoter responsive to radiation in prostate cancer cells, we prepared another promoter library consisting of 64 possible promoter fragments constructed by using the cis-acting elements of the same set of transcription factors, applying essentially the same technique, and designated it the 'four ciselement promoter library' . As many cell lines would show similar reactions to radiation, including stress responses, DNA repair and apoptosis, we assumed that a set of transcription factors activated by radiation was similar to some extent even in cell lines derived from different tissue origins.…”

“…As we reported previously, it could change the properties of a promoter to various directions. 18 Twenty-eight mutant DNA fragments were generated from clone 880 and cloned in the same luciferase expression vector as the promoter libraries. They were evaluated by dual luciferase assay 48 h after X-irradiation at 10 Gy.…”

“…However, it turned out that the response of the promoter to X-ray irradiation (X-irradiation) was cell type-dependent because no activation was observed in some of the cell lines tested. 18 In this study, we constructed a radiation-responsive promoter that functions in prostate cancer cells by using the cis-acting elements of the transcription factors activated in prostate cancer cell lines by radiation stimulation. Then we examined a constructed radiation-responsive promoter for its therapeutic use against a prostate cancer cell line.…”