Abstract
The eukaryotic translation initiation factor eIF4E acts in the nuclear export and translation of a subset of mRNAs. Both of these functions contribute to its oncogenic potential. While the biochemical mechanisms that underlie translation are relatively well understood, the molecular basis for eIF4E's role in mRNA export remains largely unexplored. To date, over 3000 transcripts, many encoding oncoproteins, were identified as potential nuclear eIF4E export targets. These target RNAs typically contain a ∼50-nucleotide eIF4E sensitivity element (4ESE) in the 3′ UTR and a 7-methylguanosine cap on the 5′ end. While eIF4E associates with the cap, an unknown factor recognizes the 4ESE element. We previously identified cofactors that functionally interacted with eIF4E in mammalian cell nuclei including the leucine-rich pentatricopeptide repeat protein LRPPRC and the export receptor CRM1/XPO1. LRPPRC simultaneously interacts with both eIF4E bound to the 5′ mRNA cap and the 4ESE element in the 3′ UTR. In this way, LRPPRC serves as a specificity factor to recruit 4ESE-containing RNAs within the nucleus. Further, we show that CRM1 directly binds LRPPRC likely acting as the export receptor for the LRPPRC-eIF4E–4ESE RNA complex. We also found that Importin 8, the nuclear importer for cap-free eIF4E, imports RNA-free LRPPRC, potentially providing both coordinated nuclear recycling of the export machinery and an important surveillance mechanism to prevent futile export cycles. Our studies provide the first biochemical framework for the eIF4E-dependent mRNA export pathway.
Highlights
The eukaryotic translation initiation factor eIF4E plays a major role in post-transcriptional regulation of gene expression (Carroll and Borden 2013). eIF4E is localized in both the nucleus and cytoplasm where it acts in the mRNA export and translation of a specific subset of mRNAs, respectively (Carroll and Borden 2013)
We examined whether LRPPRC plays a direct role in eIF4Edependent mRNA export by testing if it directly interacts with eIF4E in pull-down assays
We identify LRPPRC as the RNA specificity component of the complex, directly binding 4ESE RNAs and as the factor that interacts with the export receptor CRM1
Summary
The eukaryotic translation initiation factor eIF4E plays a major role in post-transcriptional regulation of gene expression (Carroll and Borden 2013). eIF4E is localized in both the nucleus and cytoplasm where it acts in the mRNA export and translation of a specific subset of mRNAs, respectively (Carroll and Borden 2013). The S53A eIF4E mutant does not act in mRNA export or transform cells; this mutant still binds the cap and is active in translation in the cytoplasm (Kaufman et al 1993; Cohen et al 2001; Culjkovic-Kraljacic et al 2012). To define biochemical factors in this export pathway, we carried out proteomic studies and found that the leucinerich pentatricopeptide repeat protein (LRPPRC) immunoprecipitates with both eIF4E and 4ESE RNAs in the nucleus (Topisirovic et al 2009). Knockdown of LRPPRC reduces the ability of eIF4E to immunoprecipitate with 4ESE RNA and impairs eIF4E-dependent mRNA export (Topisirovic et al 2009) These studies suggest that LRPPRC is an important factor in this pathway. Our studies provide a molecular basis for essential steps in the eIF4E-dependent mRNA export pathway as well as a molecular understanding of how cap- and RNA-binding status underlie critical control points in this process
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