Eukaryotic Initiation factor 4B is a multi-functional RNA binding protein that regulates histone mRNAs
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE233998 | GEO | 2024/09/04
REPOSITORIES: GEO
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Project description:DEAD-box RNA helicases eIF4A and Ded1 promote translation by resolving mRNA secondary structures that impede preinitiation complex (PIC) attachment to mRNA or scanning. eIF4B is a cofactor for eIF4A but might also function independently of eIF4A. Ribosome profiling of mutants lacking eIF4B or with impaired eIF4A or Ded1 activity revealed that eliminating eIF4B reduces the relative translational efficiencies of many more genes than does inactivation of eIF4A, despite comparable reductions in bulk translation, and few genes display unusually strong requirements for both factors. However, either eliminating eIF4B or inactivating eIF4A preferentially impacts mRNAs with longer, more structured 5’UTRs. These findings reveal an eIF4A-independent role for eIF4B in addition to its function as eIF4A cofactor in promoting PIC attachment or scanning on structured mRNAs. eIF4B, eIF4A, and Ded1 mutations also preferentially impair translation of longer mRNAs in a fashion mitigated by the ability to form closed-loop mRNPs via eIF4F-Pab1 association, suggesting cooperation between closed-loop assembly and eIF4B/helicase functions. Remarkably, depleting eIF4G, the scaffold subunit of eIF4F, preferentially impacts short mRNAs with strong closed-loop potential and unstructured 5’UTRs, exactly the opposite features associated with hyperdependence on the eIF4B/helicases. We propose that short, highly efficient mRNAs preferentially depend on the stimulatory effects of eIF4G-dependent closed-loop assembly.
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Project description:Cellular responses to environmental stress are frequently mediated by RNA-binding proteins (RBPs). Here, we examined global RBP dynamics in Saccharomyces cerevisiae in response to glucose starvation and heat shock. Each stress induced rapid remodeling of the RNA-protein interactome, without corresponding changes in RBP abundance. Consistent with general translation shutdown, ribosomal proteins contacting the mRNA showed decreased RNA-association. Among translation components, RNA-association was most reduced for initiation factors involved in 40S scanning (eIF4A, eIF4B, and Ded1), indicating a common mechanism of translational repression. In unstressed cells, eIF4A, eIF4B, and Ded1 primarily targeted the 5′-ends of mRNAs. Following glucose withdrawal, 5’-binding was abolished within 30sec, explaining the rapid translation shutdown, but mRNAs remained stable. Heat shock induced progressive loss of 5’ RNA-binding by initiation factors over ~16min. Translation shutoff provoked selective 5′-degradation of mRNAs encoding translation-related factors, mediated by Xrn1. These results reveal mechanisms underlying translational control of gene expression during stress.
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Project description:Rocaglamide A (RocA) typifies a class of protein synthesis inhibitors that selectively kill aneuploid tumor cells and repress translation of specific mRNAs. RocA targets eukaryotic initiation factor 4A (eIF4A), an ATP-dependent DEAD-box RNA helicase; its mRNA selectivity is proposed to reflect highly structured 5′ UTRs that depend strongly on eIF4A-mediated unwinding. However, rocaglate treatment may not phenocopy the loss of eIF4A activity, as these drugs actually increase the affinity between eIF4A and RNA. Here, we show that secondary structure in 5′ UTRs is only a minor determinant for RocA selectivity and RocA does not repress translation by reducing eIF4A availability. Rather, in vitro and in cells, RocA specifically clamps eIF4A onto polypurine sequences in an ATP-independent manner. This artificially clamped eIF4A blocks 43S scanning, leading to premature, upstream translation initiation and reducing protein expression from transcripts bearing the RocA-eIF4A target sequence. In elucidating the mechanism of selective translation repression by this lead anti-cancer compound, we provide an example of a drug stabilizing sequence-selective RNA-protein interactions.
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