Involvement of multiple factors in regulation of the CDKN1A gene promoter in response to ionizing radiation

TP53 is a tumor suppressor protein functioning as a transcription factor to regulate plenty of genes involved in physiological responses to DNA damages, such as apoptosis, cell cycle arrest, DNA repair, etc. When induction of various TP53-target genes in various organs of mouse following whole body exposure to ionizing radiation (IR) was examined, it was shown that, in a same organ, induction varies from gene to gene, and, for a same gene, induction varies from organ to organ. Selective regulation of TP53-target genes may be the basis for the multi-functional property of TP53. In this regards, it is noteworthy that some transcription factors, such as Sp1, GKLF, Ets1, and IRF-1, have been reported to play a role in TP53-dependent transcriptional activation. It may be hypothesized that variable transcription factors and cofactors participate in regulation of TP53-target genes depending on organs, their physiological conditions, radiation doses, dose-rates and so on. We here comprehensively investigated the factors involved in regulation of a TP53-target gene CDKN1A(p21) after irradiation. We first carried out an electrophoretic mobility shift assay to analyze DNA-protein interaction in the upstream regions of the CDKN1A gene using 115 probes covering the region of approximately 2.9 kb in length. A drastic change of the DNA-protein interaction was observed at -2193bp/-2185bp which is closely located to the TP53 recognition site at -2261bp. Interestingly alteration of the DNA-protein interaction at -1368bp/-1333bp and -1268bp/-1233bp was observed only after irradiation with IR in the dose range of 0.2-2 Gy. Next, we carried out a reporter gene analysis using variously deleted CDKN1A gene promoter linked to the luciferase-encoding sequence. It was found that deletion of the region -1398bp/-1119bp drastically diminished the induction of the CDKN1A gene promoter after IR in the dose range of 0.2-2 Gy. As this region contains the loci where alteration of the DNA-protein interaction occurred after IR in the dose range of 0.2-2 Gy, it was suggested that some transcription factors, inducible after IR of this dose range, bind DNA at the loci -1368bp/-1333bp and -1268bp/-1233bp to regulate the CDKN1A gene promoter in cooperation with TP53.13th International Congress of Radiation Researc