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Molecular and Cellular Biology, January 2008, p. 93-107, Vol. 28, No. 1
0270-7306/08/$08.00+0     doi:10.1128/MCB.00973-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

RNA-Binding Proteins HuR and PTB Promote the Translation of Hypoxia-Inducible Factor 1 ^,

Laboratory of Cellular and Molecular Biology, National Institute on Aging Intramural Research Program, National Institutes of Health, Baltimore, Maryland 21224,¹ Department of Pharmacology and Molecular Sciences, Johns Hopkins School of Medicine, Baltimore, Maryland 21205,² Apoptosis Research Centre, Children's Hospital of Eastern Ontario Research Institute, Ottawa, Ontario K1H 8L1, Canada³

Received 2 June 2007/ Returned for modification 16 July 2007/ Accepted 15 October 2007

The levels of hypoxia-inducible factor 1 (HIF-1) are tightly controlled. Here, we investigated the posttranscriptional regulation of HIF-1 expression in human cervical carcinoma HeLa cells responding to the hypoxia mimetic CoCl₂. Undetectable in untreated cells, HIF-1 levels increased dramatically in CoCl₂-treated cells, while HIF-1 mRNA levels were unchanged. HIF-1 translation was potently elevated by CoCl₂ treatment, as determined by de novo translation analysis and by monitoring the polysomal association of HIF-1 mRNA. An internal ribosome entry site in the HIF-1 5' untranslated region (UTR) was found to enhance translation constitutively, but it did not further induce translation in response to CoCl₂ treatment. Instead, we postulated that RNA-binding proteins HuR and PTB, previously shown to bind HIF-1 mRNA, participated in its translational upregulation after CoCl₂ treatment. Indeed, both RNA-binding proteins were found to bind HIF-1 mRNA in a CoCl₂-inducible manner as assessed by immunoprecipitation of endogenous ribonucleoprotein complexes. Using a chimeric reporter, polypyrimidine tract-binding protein (PTB) was found to bind the HIF-1 3'UTR, while HuR associated principally with the 5'UTR. Lowering PTB expression or HuR expression using RNA interference reduced HIF-1 translation and expression levels but not HIF-1 mRNA abundance. Conversely, HIF-1 expression and translation in response to CoCl₂ were markedly elevated after HuR overexpression. We propose that HuR and PTB jointly upregulate HIF-1 translation in response to CoCl₂.

Published ahead of print on 29 October 2007.

Supplemental material for this article may be found at http://mcb.asm.org/.

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