Abstract 1935: A role for microRNAs in the regulation of transcription factor ATF5

ATF5 is a widely expressed transcription factor that modulates survival, proliferation, and differentiation and plays a role in homeostasis and cellular stress response. In unstressed conditions, ATF5 has a short half-life and is rapidly degraded due to post-translational modifications. Conversely, ATF5 is upregulated in cells under cellular stress. Furthermore, we have found that ATF5 is elevated in transformed C6 glioma and MCF7 breast cancer compared to nontransformed cells and is a survival factor for both transformed cell lines. Although it is known that ATF5 mRNA can be stabilized at the 5’ untranslated region (UTR) and protected from degradation via interactions with other proteins, regulation of ATF5 expression is not fully understood. We hypothesize that microRNA (miRNA) play a role in regulating the expression of ATF5 at the 3’ UTR. MiRNAs are endogenous small non-coding RNAs 20-25 nucleotides in length that contribute to regulation of gene expression at the translational level. Herein, we sought to identify the presence of specific miRNA and their levels and ability to downregulate ATF5 during cellular stress and other physiological conditions in vitro. To date, no studies have examined the regulation of ATF5 by miRNA. Binding sites of miRNA are typically found in the UTRs of target mRNAs. We used TargetScan, miRanda, and other in silico modeling programs to identify miRNAs that were predicted to bind to the 3’ UTR of ATF5 and selected miRNAs 129-5p, 433-3p, and 520b as initial candidates for their potential in regulation of ATF5. Here, we have employed multiple strategies to investigate a potential role of miRNAs in the regulation of ATF5. Luciferase reporter assays were used as an initial method of validation for the miRNA candidates. Subsequently, transfections were carried out with precursor microRNA and expression levels of ATF5 were measured under varying physiological conditions via Western blot analysis. Additionally, immunoprecipitations were carried out with ATF5 3’ UTR mRNA and HeLa cell lysate to assess the presence of miRNAs and quantify the amount of specific miRNA bound to the 3’ UTR under varying physiological conditions via qPCR analysis. Preliminary data show that miRNA are bound to the 3’ UTR of ATF5 and that miRNAs 129-5p, 433-3p, and 520b help to regulate the expression of ATF5 under varying stress conditions and at steady state. Better understanding of the regulation of ATF5 could have implications in a broad range of human malignancies, including glioma and potentially several other types of cancer. Citation Format: Kari A. Gaither, David X. Liu, Bhanupriya Madarampalli. A role for microRNAs in the regulation of transcription factor ATF5. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1935.