Abstract 1947: Identification of microRNAs that regulate DNA damage response

Abstract

Abstract Mammalian cells have developed an elaborate DNA damage response system and DNA repair machinery, which play a critical role in the prevention of cancer and development of resistance to anti-cancer DNA damaging agents. Many proteins involved in DNA damage response, such as phosphorylated histone H2AX (γH2AX) and 53BP1, form nuclear foci at sites of DNA damage and facilitate DNA damage response and repair. The precise regulation of DNA damage response is crucial for cellular survival after DNA damage and its abrogation often results in genomic instability. MicroRNAs (miRNAs) are short, non-coding RNA molecules, which post-transcriptionally regulate gene expression by repressing translation and/or degrading mRNA. Some miRNAs function as oncogenes or tumor suppressors. However, the role of miRNAs in DNA damage response remains largely unknown. Here, we identified miRNAs that regulate DNA damage response, especially γH2AX/53BP1 foci formation, and modulate chemo-/radio-sensitivity of tumor cells. We developed a cell-based screening assay utilizing ionizing radiation (IR)-induced γH2AX/53BP1 foci formation in a human osteosarcoma cell line, U2OS, as a readout. The human miRNA Mimics and Inhibitors Libraries (Dharmacon), consisting of 810 miRNA mimics or inhibitors, were transfected in U2OS cells and screened with an automated high-throughput fluorescence microscope and an automatic foci counter. We found that two and five miRNA mimics reproducibly inhibited IR-induced γH2AX and 53BP1 foci formation, respectively, while none of the miRNA inhibitors inhibited their foci formation. Among these hits, we focused on miR-138, because miR-138 is reported to be deregulated in a subset of cancer and may be involved in tumorigenesis and chemosensitivity of cancer. We found that miR-138 decreased the expression of H2AX and reduced H2AX phosphorylation upon DNA damage. Consistent with a defect in DNA damage response, miR-138 sensitized U2OS cells to DNA damaging agents, such as IR and cisplatin. Importantly, the 3′UTR of H2AX contains a putative miR-138 binding site, suggesting that H2AX may be a direct target of miR-138. Taken together, we have identified some miRNAs which inhibit IR-induced foci formation of γH2AX/53BP1. In particular, miR-138 strongly reduced H2AX-mediated DNA damage response and enhanced the chemo-/radio-sensitivity of cancer cells, suggesting that miR-138 might serve as an important regulator of genome instability and a possible therapeutic target for cancer treatment. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1947.