Translational Dysregulation by Pateamine A (original) (raw)
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Proceedings of the National Academy of Sciences of the United States of America, 2016
DEAD-box RNA helicases eukaryotic translation initiation factor 4A (eIF4A) and Ded1 promote translation by resolving mRNA secondary structures that impede preinitiation complex (PIC) attachment to mRNA or scanning. Eukaryotic translation initiation factor 4B (eIF4B) is a cofactor for eIF4A but also might 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' untranslated regions (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,...
Journal of Biological Chemistry, 2010
Translation re-initiation provides the molecular basis for translational control of mammalian ATF4 and yeast GCN4 mediated by short upstream open reading (uORFs) in response to eIF2 phosphorylation. eIF4G is the major adaptor subunit of eIF4F that binds the cap-binding subunit eIF4E and the mRNA helicase eIF4A and is also required for re-initiation in mammals. Here we show that the yeast eIF4G2 mutations altering eIF4Eand eIF4A-binding sites increase re-initiation at GCN4 and impair recognition of the start codons of uORF1 or uORF4 located after uORF1. The increase in re-initiation at GCN4 was partially suppressed by increasing the distance between uORF1 and GCN4, suggesting that the mutations decrease the migration rate of the scanning ribosome in the GCN4 leader. Interestingly, eIF4E overexpression suppressed both the phenotypes caused by the mutation altering eIF4E-binding site. Thus, eIF4F is required for accurate AUG selection and re-initiation also in yeast, and the eIF4G interaction with the mRNA-cap appears to promote eIF4F re-acquisition by the re-initiating 40 S subunit. However, eIF4A overexpression suppressed the impaired AUG recognition but not the increase in re-initiation caused by the mutations altering eIF4A-binding site. These results not only provide evidence that mRNA unwinding by eIF4A stimulates start codon recognition, but also suggest that the eIF4A-binding site on eIF4G made of the HEAT domain stimulates the ribosomal scanning independent of eIF4A. Based on the RNA-binding activities identified within the unstructured segments flanking the eIF4G2 HEAT domain, we discuss the role of the HEAT domain in scanning beyond loading eIF4A onto the pre-initiation complex.
Molecular and Cellular Biology, 2009
Eukaryotic mRNAs possess a 5-terminal cap structure (cap), m 7 GpppN, which facilitates ribosome binding. The cap is bound by eukaryotic translation initiation factor 4F (eIF4F), which is composed of eIF4E, eIF4G, and eIF4A. eIF4E is the cap-binding subunit, eIF4A is an RNA helicase, and eIF4G is a scaffolding protein that bridges between the mRNA and ribosome. eIF4G contains an RNA-binding domain, which was suggested to stimulate eIF4E interaction with the cap in mammals. In Saccharomyces cerevisiae, however, such an effect was not observed. Here, we used recombinant proteins to reconstitute the cap binding of the mammalian eIF4E-eIF4GI complex to investigate the importance of the RNA-binding region of eIF4GI for cap interaction with eIF4E. We demonstrate that chemical cross-linking of eIF4E to the cap structure is dramatically enhanced by eIF4GI fragments possessing RNA-binding activity. Furthermore, the fusion of RNA recognition motif 1 (RRM1) of the La autoantigen to the N terminus of eIF4GI confers enhanced association between the cap structure and eIF4E. These results demonstrate that eIF4GI serves to anchor eIF4E to the mRNA and enhance its interaction with the cap structure.
The eukaryotic translation initiation factor eIF4E in the nucleus: taking the road less traveled
Immunological Reviews, 2014
The eukaryotic translation initiation factor eIF4E is a potent oncogene. Although eIF4E has traditional roles in translation initiation in the cytoplasm, it is also found in the nucleus, suggesting that it has activities beyond its role in protein synthesis. The road less traveled has been taken to study these nuclear activities and to understand their contribution to the oncogenic potential of eIF4E. The molecular features and biological pathways underpinning eIF4E's nuclear mRNA export are described. New classes of eIF4E regulators have been identified and their relevance to cancer shown. The studies presented here reveal the molecular, biophysical, and structural bases for eIF4E regulation. Finally, recent clinical work targeting eIF4E in acute myeloid leukemia patients with ribavirin is discussed. In summary, these findings provide a novel paradigm for eIF4E function and the molecular basis for targeting it in leukemia patients. Keywords eIF4E; PML; mRNA export; ribavirin; clinical trials General overview Normal development, differentiation, and cellular growth rely on post-transcriptional as well as transcriptional control of gene expression. The eukaryotic translation initiation factor, eIF4E, regulates gene expression post-transcriptionally at multiple levels including mRNA translation and mRNA export (1-4). Through these activities, eIF4E promotes cellular proliferation, growth, and survival (1-4). Even moderate overexpression of eIF4E leads to dysregulated cellular proliferation and malignant transformation. eIF4E is dysregulated in an estimated 30% of human cancers. The traditional view is that eIF4E recruits mRNAs to the ribosome and that when dysregulated, this leads to inappropriate translation leading to transformation (1). This viewpoint further suggests that eIF4E is controlled almost exclusively through the mTOR eIF4E-binding protein (BP) axis (1), in which cytoplasmic eIF4E-mediated protein synthesis is regulated by signaling events upstream of mTOR which leads to eventual phosphorylation
The eukaryotic translation initiation factor eIF4E wears a “cap” for many occasions
Translation, 2016
The eukaryotic translation initiation factor eIF4E plays important roles in controlling the composition of the proteome. Indeed, dysregulation of eIF4E is associated with poor prognosis cancers. The traditional view has been that eIF4E acts solely in translation. However, over the last »25 years, eIF4E was found in the nucleus where it acts in mRNA export and in the last »10 years, eIF4E was found in cytoplasmic processing bodies (P-bodies) where it functions in mRNA sequestration and stability. The common biochemical thread for these activities is the ability of eIF4E to bind the 7methylguanosine cap on the 5 0 end of mRNAs. Recently, the possibility that eIF4E directly binds some mRNA elements independently of the cap has also been raised. Importantly, the effects of eIF4E are not genome-wide with a subset of transcripts targeted depending on the presence of specific mRNA elements and context-dependent regulatory factors. Indeed, eIF4E governs RNA regulons through co-regulating the expression of groups of transcripts acting in the same biochemical pathways. In addition, studies over the past »15 years indicate that there are multiple strategies that regulatory factors employ to modulate eIF4E activities in context-dependent manners. This perspective focuses on these new findings and incorporates them into a broader model for eIF4E function.
The EMBO Journal
Communicated by J.Schlessinger Eukaryotic translation initiation factor4A (eLF4A) plays a critical role in binding of eukaryotic mRNAs to ribosomes. It has been biochemically characterized as an RNA-dependent ATPase and RNA helicase and is a prototype for a growing family of putative RNA helicases termed the DEAD box family. It is required for mRNA-ribosome binding both in its free form and as a subunit of the cap binding protein complex, eIF-4F. To gain further understanding into the mechanism of action of eIF-4A in mRNA-ribosome binding, defective eIF-4A mutants were tested for their abilities to function in a dominant negative manner in a rabbit reticulocyte translation system. Several mutants were demonstrated to be potent inhibitors of translation. Addition of mutant eI]F4A to a rabbit reticulocyte translation system strongly inhibited translation of all mRNAs studied including those translated by a cap-independent internal initiation mechanism. Addition of eIF4A or eIF-4F relieved inhibition of translation, but eIF-4F was six times more effective than eIF-4A, whereas eIF-4B or other translation factors failed to relieve the inhibition. Kinetic experiments demonstrated that mutant eIF-4A is defective in recycling through eIF-4F, thus explaining the dramatic inhibition of translation. Mutant eIF-4A proteins also inhibited eEF-4F-dependent, but not eIF-4Adependent RNA helicase activity. Taken together these results suggest that eIF-4A functions primarily as a subunit of eIF-4F, and that singular eIF-4A is required to recycle through the complex during translation. Surprisingly, eIF-4F, which binds to the cap structure, appears to be also required for the translation of naturally uncapped mRNAs.
eIF4G - An integrator of mRNA metabolism?
FEMS yeast research, 2016
The eukaryotic translation initiation factor, eIF4G plays a key functional role in the initiation of cap-dependent translation by acting as an adapter to nucleate the assembly of eIF4F complex. Together with poly(A) binding protein and eIF3, eIF4F subsequently triggers the recruitment of 43S ribosomal pre-initiation complex to the messenger RNA template. Since eukaryotes primarily regulate translation at the level of initiation, eIF4G is implicated in the control of eukaryotic gene expression. Remarkably, emerging evidence in Saccharomyces cerevisiae indicates that eIF4G also plays a key role in nuclear mRNA biogenesis and surveillance - a finding that is in agreement with its nuclear distribution. Here, we focus on the functional involvement of eIF4G in the nucleus in modulating pre-mRNA splicing, mRNA surveillance, and possibly in much-debated nuclear translation. Notably, the nature of the biochemical role of this protein in the major events of cellular mRNA metabolism emphasizes...
Structural Studies on the eIF4A-eIF4G Interaction in Translation Initiation
2012
Protein synthesis is an important cellular process, and the RNA helicase eIF4A plays a vital role in unwinding messenger RNA and scanning during translation initiation. eIF4A has little activity in isolation, but is modulated by other initiation factors such as eIF4G and eIF4H. In this thesis, we explore how these proteins come together to form a functional unwinding complex. We begin with the NMR solution structure of a single domain from this complex, eIF4G HEAT2. We then map interactions involving HEAT2 and its binding partners, as well as those involving the N-terminal domain of eIF4A. We use this information first to construct a structure of the two-domain complex of HEAT2 and eIF4A-NTD, and expand this work toward the structure of the 70kDa, three-domain complex of HEAT2 with full-length eIF4A. Finally, we incorporate eIF4H and another domain of eIF4G to model the entire functional complex, and explore how interactions between domains rearrange upon binding, hydrolysis, and release of ATP. These results give us a better understanding of how eIF4G modulates eIF4A helicase activity. Moreover, the domain organization of the complex allows us to construct a more compelling model to explain how eIF4A facilitates preinitiation complex scanning along a messenger RNA.