Probing the closed-loop model of mRNA translation in living cells.
The mRNA closed-loop, formed through interactions between the cap structure, poly(A) tail, eIF4E, eIF4G and PAB, features centrally in models of eukaryotic translation initiation, although direct support for its existence in vivo is not well established. Here, we investigated the closed-loop using a combination of mRNP isolation from rapidly cross-linked ... cells and high-throughput qPCR. Using the interaction between these factors and the opposing ends of mRNAs as a proxy for the closed-loop, we provide evidence that it is prevalent for eIF4E/4G-bound but unexpectedly sparse for PAB1-bound mRNAs, suggesting it primarily occurs during a distinct phase of polysome assembly. We observed mRNA-specific variation in the extent of closed-loop formation, consistent with a role for polysome topology in the control of gene expression.
Mesh Terms:
Binding Sites, Eukaryotic Initiation Factor-4E, Eukaryotic Initiation Factor-4G, Gene Expression Regulation, Fungal, Nucleic Acid Conformation, Poly(A)-Binding Proteins, Polyribosomes, Protein Binding, Protein Biosynthesis, RNA, Fungal, RNA, Messenger, Recombinant Proteins, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins
Binding Sites, Eukaryotic Initiation Factor-4E, Eukaryotic Initiation Factor-4G, Gene Expression Regulation, Fungal, Nucleic Acid Conformation, Poly(A)-Binding Proteins, Polyribosomes, Protein Binding, Protein Biosynthesis, RNA, Fungal, RNA, Messenger, Recombinant Proteins, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins
RNA Biol
Date: Apr. 01, 2015
PubMed ID: 25826658
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