Browsing by Issue Date, starting with "2021-05-25"
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- Human TP53 is a tumour suppressor that acquires oncogenic functions during integrated stress response (ISR) due to a translational switchPublication . Lacerda, Rafaela; Fonseca Costa, Inês; López-Iniesta, Maria; Romão, Luísa; Candeias, Marco M.Eukaryotic cells have developed different mechanisms and adaptive pathways that allow them to cope with external stress stimuli. Under stress conditions, global protein synthesis is shut down, and some alternative mechanisms of mRNA translation initiation are induced. Although the tumour suppressor protein p53 — the most mutated gene in cancer — has been considered the guardian of the genome and a master regulator of several cellular functions, the truth is it is not just one isoform, the full-length (FLp53), but also many other p53 isoforms that have been described so far. Based on our previous results, some functions of the shorter isoforms are different from and complement FLp53 activity. Here we show the specific induction of Δ160p53 isoform during integrated stress response (ISR). We confirmed the presence of an Internal Ribosome Entry Site (IRES) in p53 mRNA that controls Δ160p53 isoform translation, using a bicistronic reporter construct. When subjecting cells to endoplasmic reticulum stress, we showed that eIF2α phosphorylation is a key event leading to cap-independent expression of Δ160p53 during ISR. Also, some cancer-specific mutations in the DNA-binding domain of p53 enhance cap-independent translation of Δ160p53 via Δ160p53IRES. Using an antisense morpholino oligo targeting Δ160IRES significantly reduces Δ160p53 protein levels and impaired its oncogenic functions. Additionally, we found the 5’untranslated region of Δ160p53 inhibits the IRES activity. Our data support a model in which an IRES structure in the coding region of p53 is activated under stress conditions, leading to the expression of the oncogenic shorter Δ160p53 isoform, whose structure is affected by cancer-specific mutations in the p53 gene. A better understanding of Δ160p53IRES structure and function may be advantageous for a more efficient therapeutic targeting of p53.
- Regulation of the Human PERK mRNA Translation by Upstream Open Reading FramesPublication . Fernandes, Rafael; Rodrigues, Rosário; Lopes, Pedro; Romão, LuísaUpstream open reading frames (uORFs) are cis-acting elements located within the 5’ leader sequence (5’UTR) of transcripts, which can regulate translation of the correspondent main open reading frame (mORF). During endoplasmic reticulum (ER) stress, the accumulation of unfolded proteins activates the ER-resident PKR-like ER kinase (PERK), which results in phosphorylation of eIF2α to inhibit global mRNA translation, while allowing the selective uORF-mediated translation of downstream effectors responsible for stress resolution or, ultimately, cell death. The dual role of PERK in regulating cell fate was implicated in human diseases, like diabetes, neurodegenerative disorders and cancer. Moreover, mutations in the EIF2AK3 gene (encoding PERK) were associated to the rare genetic disease, Wolcott-Rallison Syndrome (WRS). In this work, we aimed to study the translational regulatory role of 5 AUG- and 3 non-AUG-uORFs identified in the PERK 5’UTR and assess its biological relevance. While uORF2 and the non-AUG-uORFs 5, 6 and 7 (numbered according to their distance to the 5’ end of the mRNA) do not seem to have a regulatory role, uORF1, uORF3, uORF4 and uORF8 together present a strong repressive effect over mORF translation in basal conditions. Also, we observe that uORF1 is frequently translated allowing low levels of translation re-initiation at the main ORF. Curiously, we found that when PERK is overexpressed, it leads to the spontaneous activation of a portion of PERK in the absence of any stress stimulus, possibly highlighting the biological relevance of its uORF-mediated translational regulation. Conversely, during ER stress, increased bypass of uORF1 results in a modest degree of translational de-repression, which may help to counterbalance the increased rate of PERK protein turnover observed in these conditions. We also observed that alteration of the PERK uORFs by mutations found in WRS patients modify mORF expression, providing a possible link to the disease. Altogether, we highlight the importance of including 5’UTRs in the screening of disease-related mutations and the necessity of functional studies to assess their role in pathogenesis.
- Contaminantes Metálicos nos Alimentos - Aula teóricaPublication . Santiago, Susana; Nascimento, AnaPalestra sobre contaminantes metálicos nos alimentos.
- Contaminantes Metálicos nos Alimentos - Aula teórico-práticaPublication . Nascimento, Ana; Santiago, SusanaPalestra sobre contaminantes metálicos nos alimentos.