Abstract 2125: Cis-mediated regulation of mRNA translation initiation of p53 family members

Abstract

Abstract The p53 family of transcription factors comprises several proteins produced by three genes, TP63, TP63 and TP73, through alternative promoters and splicing, that exhibit critical functions for cell homeostasis and act as prominent tumor suppressors. The p53 pathway is often activated in stress conditions during which the global cap-dependent translation is inhibited. Although p53 protein is highly regulated at post-translational level, cells maintain continuous p53 protein synthesis also taking advantage of the presence of two Internal Ribosome Entry Sites (IRESs) elements residing within the p53 mRNA. The first IRES is located in the 5′UTR of the full-length isoform, the second is located into the protein-coding region and mediates the translation of a ΔN-p53 isoform. IRES elements are complex RNA structural elements in 5′UTRs that can mediate cap-independent initiation of translation under stress conditions. Even though an IRES activity has been already established for p53, no studies are available for p63 and p73 transcripts. This work is mainly focused on the regulation of translation initiation efficiency for the p53 family members p63 and p73. For this purpose, we have cloned the 5′UTR from human full-length p53 (as positive control), TA-p63, TA*-p63, TA-p73 and ΔN-p63 and ΔN-p73 transcript isoforms in a bicistronic reporter construct (named pRuF, where R stands for Renilla reniformis luciferase cDNA, u for 5′UTR tested and F for Firefly luciferase cDNA). The c-Myc and p16 or β-globin and β-actin 5′UTRs were included as positive or negative controls. We performed gene reporter assays in MCF7 and HCT116 p53 wild-type cancer cell lines and in their derivative clones with reduced (shp53) or null (p53-/-) p53 expression. It has been previously demonstrated that p53, through transcriptional repression of fibrillarin (FBL) is able to negatively impact on rRNA methylation pattern and IRES-dependent translation, hence p53 status can be an important factor to unmask cap-independent translation initiation. Results showed that TA*-p63, TA-p73 and ΔN-p73 can potentially contain an IRES-like sequence in their 5′UTR. Translation stimulation was increased in MCF7 cells with p53 was knocked-down. Treatment with the mTOR inhibitors Rapamycin and Torin1, that inhibit cap-dependent translation, led to an enhanced relative activity of the putative IRES contained in TA-p73. Interestingly, differences in responses of the various 5′UTRs between the two cell lines were observed. Based on these initial observations, the identification of different and tissue specific RNA binding proteins targeting the distinct p63 and p73 5′UTRs will be pursued with the aim to dissect a mechanism regulating mRNA translation initiation that could play an important role in development as well as in cancer progression. Note: This abstract was not presented at the meeting. Citation Format: Alessandra Bisio, Alberto Inga. Cis-mediated regulation of mRNA translation initiation of p53 family members. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2125. doi:10.1158/1538-7445.AM2015-2125