Involvement of Oct-1 in the regulation of CDKN1A in response to clinically relevant doses of ionizing radiation.

ABSTRACT

CDKN1A is a cyclin-dependent kinase inhibitor that plays a critical role in cell cycle checkpoint regulation. It is transcriptionally induced by TP53 (p53) following exposure to ionizing radiation (IR). Induction of CDKN1A after irradiation is closely related to IR-sensitivity of tumor cells, but the underlying mechanisms remain obscure because conventional reporter gene systems respond poorly to IR unless hyperlethal doses are used. Here, we performed a promoter analysis of the CDKN1A gene following irradiation with clinically relevant doses of IR using the adeno-associated virus-mediated reporter system which we have recently shown to be highly responsive to IR. We demonstrate that there are regulatory elements at -1.1 kb, -1.4 kb, and -1.8 kb, and deletion of these elements attenuate induction of the CDKN1A gene promoter in response to 0.2-2.0 Gy of IR. EMSA and ChIP assays showed that Oct-1 binds constitutively to the elements at -1.1 kb and -1.8 kb. Functional involvement of Oct-1 was confirmed by RNA interference targeting the Oct-1 gene, which suppressed both the basal and IR-inducible components of the CDKN1A expression. Thus, our results reveal that Oct-1 is crucial to the TP53-mediated regulation of the CDKN1A gene promoter following exposure to clinically relevant doses of IR.