Translational Control of Δ160p53 Keeps the Dark Side of TP53 in Check

Ramalho, Ana Catarina; Lacerda, Rafaela; Romão, Luísa; Candeias, Marco M.

http://hdl.handle.net/10400.18/8919

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Authors

Ramalho, Ana Catarina

Lacerda, Rafaela

Romão, Luísa

Candeias, Marco M.

Abstract(s)

The TP53 tumour suppressor gene was discovered over 40 years ago, but to this day some aspects of its regulation and function remain a mystery. It encodes the full-length p53 protein (FLp53), a transcription factor with a key role in stress response in multicellular organisms, that can either direct cells towards apoptosis or recovery of homeostasis. With such a decisive role, its expression and activity are tightly regulated. A vast set of RNA-binding proteins (RBPs) have been described to affect the translation of FLp53 or the stability of p53 mRNA in response to different perturbations. But in addition to FLp53, there is a group of shorter protein isoforms lacking the N-terminal region, which have well-described functions, and are translated from the same mRNA. The shorter and less studied isoform is Δ160p53, which promotes cell survival, proliferation, and invasion. Despite its usual low levels, it is commonly overexpressed in tumours. However, the detailed mechanisms and factors involved in the regulation of Δ160p53 are still unknown. In this work, a mass spectrometry was performed to identify the proteins in an RNA-protein co-immunoprecipitation of the p53 mRNA using the MS2 system in the p53-null cell line H1299. The validation of the hits was undertaken by western blot with specific antibodies after immunoprecipitation. The effect of the binding proteins on the translation of Δ160p53 was assessed by overexpression or knockdown, and the expression levels were verified by western blot or luminescence assays. The mass spectrometry allowed the identification of potential new binding partners of the p53 mRNA. Resorting to the literature and to computational tools available online to predict protein-RNA interactions, a few hits were selected for follow-up and their interactions confirmed. Simultaneously, the modulation of Δ160p53 expression by some of these proteins was verified. Considering the importance of TP53 in deciding the fate of the cell, the observation of abnormal levels of the oncoprotein Δ160p53 in cancer is intriguing. Understanding the control of its translation could uncover strategies to block it and pave way for new cancer therapies.