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

Abstract

The type I insulin-like growth factor receptor (IGF-IR) is an integral component in the control of cell proliferation, differentiation and apoptosis. The IGF-IR mRNA contains an extraordinarily long (1038 nt) 5′-untranslated region (5′-UTR), and we have characterized a diverse series of proteins interacting with this RNA sequence which may provide for intricate regulation of IGF-IR gene expression at the translational level. Here, we report the purification and identification of one of these IGF-IR 5′-UTR-binding proteins as HuR, using a novel RNA crosslinking/RNase elution strategy. Because HuR has been predominantly characterized as a 3′-UTR-binding protein, enhancing mRNA stability and generally increasing gene expression, we sought to determine whether HuR might serve a different function in the context of its binding the IGF-IR 5′-UTR. We found that HuR consistently repressed translation initiation through the IGF-IR 5′-UTR. The inhibition of translation by HuR was concentration dependent, and could be reversed in trans by addition of a fragment of the IGF-IR 5′-UTR containing the HuR binding sites as a specific competitor, or abrogated by deletion of the third RNA recognition motif of HuR. We determined that HuR repressed translation initiation through the IGF-IR 5′-UTR in cells as well, and that siRNA knockdown of HuR markedly increased IGF-IR protein levels. Interestingly, we also found that HuR potently inhibited IGF-IR translation mediated through internal ribosome entry. Kinetic assays were performed to investigate the mechanism of translation repression by HuR and the dynamic interplay between HuR and the translation apparatus. We found that HuR, occupying a cap-distal position, significantly delayed translation initiation mediated by cap-dependent scanning, but was eventually displaced from its binding site, directly or indirectly, as a consequence of ribosomal scanning. However, HuR perpetually blocked the activity of the IGF-IR IRES, apparently arresting the IRES-associated translation pre-initiation complex in an inactive state. This function of HuR as a 5′-UTR-binding protein and dual-purpose translation repressor may be critical for the precise regulation of IGF-IR expression essential to normal cellular homeostasis.

MicroRNA-214 Reduces Insulin-like Growth Factor-1 (IGF-1) Receptor Expression and Downstream mTORC1 Signaling in Renal Carcinoma Cells

“…Translational regulation of IGF-1R is also reported. The long 5ЈUTR of IGF-1R is a target of the RNA-binding protein HuR, which delays cap-dependent translation and inhibits the internal ribosome entry site-containing IGF-1R translational block (63,64). In contrast, heterogeneous nuclear ribonucleoprotein C has been shown to interact with IGF-1R 5ЈUTR at the site of HuR binding to promote internal ribosome entry site-mediated translation of IGF-1R (65).…”

Translational Control of Cytochrome c by RNA-Binding Proteins TIA-1 and HuR

“…HuR has been shown to promote the translation of several target mRNAs with which it complexes via their 3ЈUTR (30, 35) but appears to suppress the translation of mRNAs exhibiting 5ЈUTR associations (28,37). While these discrepant roles for HuR remain to be formally examined, the data obtained here support a role for HuR in promoting cytochrome c translation under unstressed conditions: (i) binding of HuR to the cytochrome c mRNA was more abundant in unstressed cells, decreasing when translation was lowered by Tn treatment (Fig.…”

Differential NKp30 Inducibility in Chimpanzee NK Cells and Conserved NK Cell Phenotype and Function in Long-Term HIV-1-Infected Animals

“…The specific interactions between NKp30 and its ligand(s) expressed on DCs leads to NK cell activation, immature DC maturation, and immature DC killing by NK cells in humans (37,54). Little or nothing is known as of the nature of the interaction between these cells in P. troglodytes.…”