Two internal ribosome entry sites mediate the translation of p53 isoforms

Ray, Partho Sarothi; Grover, Richa; Das, Saumitra

doi:10.1038/sj.embor.7400623

Cited by 181 publications

(236 citation statements)

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“…p53-null H1299 cells were transfected with luciferase bicistronic constructs containing p53 1-251 RNA in the intercistronic region. 7,8,11 Control cells and glucose-starved cells were harvested 4, 8, 20 and 30 h post-transfection. There was a consistent increase in the relative IRES activity at all these time points following glucose deprivation, with a 1.7-fold increase in activity by 20 h (Figure 1a, H1299 panel and Supplementary Table S1).…”

Section: Resultsmentioning

confidence: 99%

“…Dual-luciferase constructs of the p53 (1-251) IRES, first (1-134) IRES, second (135-251) IRESs, 3,8 VEGF and HIF1α IRESs (kind gifts from Professor Greg Goodall, Centre of Cancer Biology, Australia and Professor Annapoorni Rangarajan, MRDG, IISc) and HCV IRES 62 were used in quantitative bicistronic assays. WWP luciferase (a kind gift from Professor Kumaravel Somasundaram, IISc) was used for measuring p53-dependent transactivation, CMV-Renilla-luciferase (Promega, Madison, WI, USA) constructs were used to normalize transfection efficiency.…”

Section: Methodsmentioning

confidence: 99%

“…[2][3][4][5][6][7] Thus, p53 mRNA has a dual IRES structure. 8 For their function, these IRESs rely on IRES trans-acting factors (ITAFs). Polypyrimidine-tract-binding protein (PTB) was shown to have differential affinity for the two IRESs of p53 and regulated p53 IRES functions by translocating from nucleus to cytoplasm on doxorubicin-induced DNA damage.…”

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Reversible induction of translational isoforms of p53 in glucose deprivation

et al. 2015

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Tumor suppressor protein p53 is a master transcription regulator, indispensable for controlling several cellular pathways. Earlier work in our laboratory led to the identification of dual internal ribosome entry site (IRES) structure of p53 mRNA that regulates translation of full-length p53 and Δ40p53. IRES-mediated translation of both isoforms is enhanced under different stress conditions that induce DNA damage, ionizing radiation and endoplasmic reticulum stress, oncogene-induced senescence and cancer. In this study, we addressed nutrient-mediated translational regulation of p53 mRNA using glucose depletion. In cell lines, this nutrient-depletion stress relatively induced p53 IRES activities from bicistronic reporter constructs with concomitant increase in levels of p53 isoforms. Surprisingly, we found scaffold/matrix attachment region-binding protein 1 (SMAR1), a predominantly nuclear protein is abundant in the cytoplasm under glucose deprivation. Importantly under these conditions polypyrimidine-tractbinding protein, an established p53 ITAF did not show nuclear-cytoplasmic relocalization highlighting the novelty of SMAR1-mediated control in stress. In vivo studies in mice revealed starvation-induced increase in SMAR1, p53 and Δ40p53 levels that was reversible on dietary replenishment. SMAR1 associated with p53 IRES sequences ex vivo, with an increase in interaction on glucose starvation. RNAi-mediated-transient SMAR1 knockdown decreased p53 IRES activities in normal conditions and under glucose deprivation, this being reflected in changes in mRNAs in the p53 and Δ40p53 target genes involved in cell-cycle arrest, metabolism and apoptosis such as p21, TIGAR and Bax. This study provides a new physiological insight into the regulation of this critical tumor suppressor in nutrient starvation, also suggesting important functions of the p53 isoforms in these conditions as evident from the downstream transcriptional target activation. Cell Death and Differentiation (2015) 22, 1203-1218; doi:10.1038/cdd.2014.220; published online 27 February 2015 p53 is a master transcription factor and tumor suppressor. Apart from post-translational modifications of p53 and concomitant protein-protein interactions of p53 with diverse factors, translational control of p53 mRNA also plays an important role under stress conditions. 1 p53 and its N-terminally truncated isoform Δ40p53 (also known as ΔN-p53 or p53/47) are translated by internal ribosome entry site (IRES)-mediated translation initiation from the same mRNA under different stress conditions that induce DNA damage, ionizing radiation and endoplasmic reticulum (ER) stress, oncogene-induced senescence and cancer. [2][3][4][5][6][7] Thus, p53 mRNA has a dual IRES structure. 8 For their function, these IRESs rely on IRES trans-acting factors (ITAFs). Polypyrimidine-tract-binding protein (PTB) was shown to have differential affinity for the two IRESs of p53 and regulated p53 IRES functions by translocating from nucleus to cytoplasm on doxorubicin-induced DNA damage. 3 This PTB-med...

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Reversible induction of translational isoforms of p53 in glucose deprivation

et al. 2015

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“…Insertion of an open-reading frame followed by a hairpin structure in the 5 0 -untranslated region of p53 resulted in an increase in p53/47 synthesis and a decrease in p53, indicating that cap-independent mechanisms of initiation control p53/47 translation via an internal ribosome entry site (IRES) structure located in the region of þ 1 to þ 120 of p53. Hence, the coding region of p53 serves as IRES for p53/47 (Candeias et al, 2006;Ray et al, 2006;Grover et al, 2009). It has also been shown that p53/47 could be generated by alternative splicing (Ghosh et al, 2004), but it should be a rare event as the full-length p53 mRNA was evident after 20 quantitative PCR cycles, whereas the p53/47 message required 40 quantitative PCR cycles, which represents approximately 2 Â 10 20 times less amount of p53/47 mRNA.…”

Section: Introductionmentioning

confidence: 99%

p53 isoforms gain functions

Olivares‐Illana

Fåhræus

2010

Oncogene

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Many different cell stress pathways converge on p53 to induce a number of distinct cell biological responses such as G1 or G2 arrest, senescence or apoptosis. One of the outstanding questions with regard to p53 is how the cells can differentiate between different stresses so that p53 activation leads to the correct response. It has been known for some time that the p53 gene expresses isoforms that carry unique domains and properties, and more recent works have started to reveal some of their functions. The alternative mRNA translation product p53/47, which lacks the first 40 codons, including the first of p53's two trans-activation domains, is being linked to endoplasmic reticulum stress and the unfolded protein response to which it causes a specific G2 arrest. On the other hand, p53 itself induces G1 arrest and has no effect on the G2. The two isoforms D133p53, which lacks the first 133 amino acids, and p53b, which carries an alternative C-terminus, are derived from alternative promoter usage or splicing, respectively, and are together implied in controlling cellular senescence. Hence, through different mechanisms of gene expression control, alternative levels of p53 isoforms help the cell to differentiate between p53 activation and the response to diverse stresses. This holds promise to a better understanding of how upstream and downstream p53 pathways have evolved relative to specific p53 domains.

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“…The manner in which this 'forced' isoform translation takes place owing to the 5 0 nonsense mutations is unknown and needs further investigation. Previous reports have shown that internal ribosomal entry sites can play a role and can be phase dependent on the cell cycle (Ray et al, 2006b;Grover et al, 2009). Another option might be that translational leakage occurs and 'false' initiation takes place at codon 41, which is surrounded by a Kozak consensus sequence (Kozak, 1984).…”

Section: Discussionmentioning

confidence: 99%

Genotoxic exposure: novel cause of selection for a functional ΔN-p53 isoform

Melis¹,

Hoogervorst²,

Oostrom³

et al. 2010

Oncogene

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The p53 gene is frequently mutated in cancers and it is vital for cell cycle control, homeostasis and carcinogenesis. We describe a novel p53 mutational spectrum, different to those generally observed in human and murine tumors. Our study shows a high prevalence of nonsense mutations in the p53 N terminus of 2-acetylaminofluorene (2-AAF)-induced urinary bladder tumors. These nonsense mutations forced downstream translation initiation at codon 41 of Trp53, resulting in the aberrant expression of the p53 isoform DN-p53 (or p44). We propose a novel mechanism for the origination and the selection for this isoform. We show that chemical exposure can act as a novel cause of selection for this truncated protein. In addition, our data suggest that the occurrence of DN-p53 accounts, at least in mice, for a cancer phenotype. We also show that gene expression profiles of embryonic stem (ES) cells carrying the DN-p53 isoform in a p53-null background are divergent from p53 knockout ES cells, and therefore postulate that DN-p53 itself has functional transcriptional properties.

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Two internal ribosome entry sites mediate the translation of p53 isoforms

Cited by 181 publications

References 23 publications

Reversible induction of translational isoforms of p53 in glucose deprivation

Reversible induction of translational isoforms of p53 in glucose deprivation

p53 isoforms gain functions

Genotoxic exposure: novel cause of selection for a functional ΔN-p53 isoform

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