The 890–1038 nt fragment of the IGF-IR 5′-UTR, containing the HuR binding sites and internal ribosome entry window, accurately recapitulates cap-dependent and IRES-mediated translation initiation and HuR repression

<p><b>Copyright information:</b></p><p>Taken from "The ELAV RNA-stability factor HuR binds the 5′-untranslated region of the human IGF-IR transcript and differentially represses cap-dependent and IRES-mediated translation"</p><p>Nucleic Acids Research 2005;33(9):2962-2979.</p><p>Published online 24 May 2005</p><p>PMCID:PMC1140080.</p><p>© The Author 2005. Published by Oxford University Press. All rights reserved</p> () translation of the control RLuc ( luciferase), full-length IGFIR (1–1038)-FLuc, and IGFIR(890–1038)-FLuc reporter RNA (each capped, 5 ng) was performed in the absence or presence of 1 or 3 mM cap analog. Each of the reporter RNAs is diagrammed schematically at the top. The luciferase activities are expressed as a percentage of the control reactions without cap-analog. () Purified recombinant GST-HuR (62 kDa, 10 ng) was incubated with the [P]UTP-labeled IGFIR(890–1038)-FLuc, FLuc or RLuc reporter RNAs, and UV crosslinking was performed as described in Materials and Methods. Samples were resolved on 10% SDS–PAGE and visualized by autoradiography. The position of the crosslinked GST-HuR band is indicated by an arrow. () translation of the IGFIR(890–1038)-FLuc reporter RNA (5 ng) with the internal control RLuc RNA (1 ng) was performed with increasing concentrations of GST-HuR (or GST as a control). The amount of HuR included in each reaction is indicated. The raw data for both the IGFIR(890–1038)-FLuc and the internal control RLuc RNAs are shown. The luciferase activities are expressed relative to the control reaction without GST-HuR. These experiments were repeated at least three times