RNA-binding protein HuR enhances p53 translation in response to ultraviolet light irradiation

Duke University, Durham, North Carolina, United States
Proceedings of the National Academy of Sciences (Impact Factor: 9.81). 08/2003; 100(14):8354-9. DOI: 10.1073/pnas.1432104100
Source: PubMed

ABSTRACT Exposure to short-wavelength UV light (UVC) strongly induces p53 expression. In human RKO colorectal carcinoma cells, this increase was not due to elevated p53 mRNA abundance, cytoplasmic export of p53 mRNA, or UVC-triggered stabilization of the p53 protein. Instead, p53 translation was potently enhanced after UVC irradiation. The 3' UTR of p53 was found to be a target of the RNA-binding protein HuR in a UVC-dependent manner in vitro and in vivo. HuR-overexpressing RKO cells displayed elevated p53 levels, whereas cells expressing reduced HuR showed markedly diminished p53 abundance and p53 translation. Our results demonstrate a role for HuR in binding to the p53 mRNA and enhancing its translation.

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    ABSTRACT: ABSTRACT Human antigen R (HuR) is a 32 kDa protein with three RNA Recognition Motifs (RRMs), which bind to Adenylate and uridylate Rich Elements (AREs) of messenger RNAs. Whereas the N-terminal and central domains (RRM1 and RRM2) are essential for AREs recognition, little is known on the C-terminal RRM3 beyond its implication in HuR oligomerization and apoptotic signaling. We have developed a detergent-based strategy to produce soluble RRM3 for structural studies. We have found that it adopts the typical RRM fold, does not interact with the RRM1 and RRM2 modules, and forms dimers in solution. Our NMR measurements, combined with Molecular Dynamics simulations and Analytical Ultracentrifugation experiments, show that the protein dimerizes through a helical region that contains the conserved W261 residue. We found that HuR RRM3 binds to 5´-mer U-rich RNA stretches through the solvent exposed side of its β-sheet, located opposite to the dimerization site. Upon mimicking phosphorylation by the S318D replacement, RRM3 mutant shows less ability to recognize RNA due to an electrostatic repulsion effect with the phosphate groups. Our study brings new insights of HuR RRM3 as a domain involved in protein oligomerization and RNA interaction, both functions regulated by two surfaces on opposite sides of the RRM domain.
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    ABSTRACT: Post-transcriptional processing of mRNA transcripts plays a critical role in establishing the gene expression profile of a cell. Such processing events are mediated by a host of factors, including RNA binding proteins and miRNAs. A number of critical cellular pathways are subject to regulation at multiple levels that allow fine-tuning of key biological responses. Programmed Cell Death 4 (PDCD4) is a tumor suppressor and an important modulator of mRNA translation that is regulated by a number of mechanisms, most notably as a target of the oncomiR, miR-21. Here we provide evidence for post-transcriptional regulation of PDCD4 by the RNA binding proteins, HuR and TIA1. Complementary approaches reveal binding of both HuR and TIA1 to the PDCD4 transcript. Consistent with a model where RNA binding proteins modulate the PDCD4 transcript, knockdown of HuR and/or TIA1 results in a significant decrease in steady-state PDCD4 mRNA and protein levels. However, fractionation experiments suggest that the mode of regulation of the PDCD4 transcript likely differs in the cytoplasm and the nucleus as the pool of PDCD4 mRNA present in the cytoplasm is more stable than the nuclear pool of PDCD4 transcript. We observe a competitive mode of binding between HuR and TIA1 on the PDCD4 transcript in the cytoplasm, suggesting that these two factors dynamically interact with one another as well as the PDCD4 transcript to maintain tight control of PDCD4 levels. Overall, this study reveals an additional set of regulatory interactions that modulate the expression of PDCD4, a key pro-apoptotic factor, and also reveal new insights into how HuR and TIA1 functions are integrated to achieve such regulation. Copyright © 2014, The American Society for Biochemistry and Molecular Biology.
    Journal of Biological Chemistry 12/2014; DOI:10.1074/jbc.M114.631937 · 4.60 Impact Factor
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