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Biochem Biophys Res Commun. 2009 Aug 14;386(1):82-8. doi: 10.1016/j.bbrc.2009.05.146. Epub 2009 Jun 6.

Fine mapping of interactions between eEF1alpha protein and 3'UTR of metallothionein-1 mRNA.

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Institute for Cell and Molecular Biosciences, Newcastle University, The Medical School, Framlington Place, Newcastle upon Tyne NE2 4HH, UK.


The localization of metallothionein-1 (MT-1) mRNA to the perinuclear cytoskeleton is determined by a signal in the 3'untranslated region (3'UTR) and trans-acting binding proteins. The present study carried out detailed mapping of this signal and further characterized the binding to elongation factor 1 alpha (eEF1alpha) and other interacting proteins. Electrophoresis mobility shift assays demonstrated that shortening of a stem region proximal to nucleotides 66-76 abrogated binding. Full length recombinant rat eEF1alpha, and independently domains I and III, formed complexes with the mRNA. Proteins binding to biotinylated MT-1 3'UTR sequences were isolated using RNA-affinity techniques, and mass spectrometry identified histidine-tRNA ligase as one of the major MT-1 3'UTR binding proteins. We conclude that a 5-bp internal stem in the MT-1 3'UTR is critical for binding of eEF1alpha and histidine-tRNA ligase, and that binding of eEF1alpha is facilitated through domains I and III.

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