EIF4A

We have developed a deep sequencing-based approach, Rec-Seq, that allows simultaneous monitoring of ribosomal 48S preinitiation complex (PIC) formation on every mRNA in the translatome in an in vitro reconstituted system. Rec-Seq isolates key early steps in translation initiation in the absence of all other cellular components and processes. Using this approach, we show that the DEAD-box ATPase Ded1 promotes 48S PIC formation on the start codons of >1000 native mRNAs, most of which have long, structured 5'-untranslated regions (5'UTRs)...

Messenger RNA (mRNA) recruitment to the 40S ribosomal subunit is mediated by eukaryotic initiation factor 4F (eIF4F). This complex includes 3 subunits: eIF4E (m7 G cap binding protein), eIF4A (DEAD-box helicase), and eIF4G. Mammalian eIF4G is a scaffold that coordinates the activities of eIF4E and eIF4A and provides a bridge to connect the mRNA and 40S ribosomal subunit through its interaction with eIF3. While the roles of many eIF4G binding domains are relatively clear, the precise function of RNA binding by eIF4G remains to be elucidated...

As obligate parasites, viruses have evolved multiple strategies to evade the host immune defense. Manipulation of the host proteasome system to degrade specific detrimental factors is a common viral countermeasure. To identify host proteins targeted for proteasomal degradation by porcine reproductive and respiratory syndrome virus (PRRSV), we conducted a quantitative proteomics screen of PRRSV-infected Marc-145 cells under the treatment with proteasome inhibitor MG132. The data revealed that the expression levels of programmed cell death 4 (PDCD4) were strongly downregulated by PRRSV and significantly rescued by MG132...

ETHNOPHARMACOLOGICAL RELEVANCE: Huangqin Qingre Chubi Capsule (HQC) is a Chinese medicinal compound used for the treatment of damp-heat pattern rheumatism, guided by the traditional Chinese medicine syndrome differentiation practice. HQC has been used in the clinical treatment of rheumatic diseases for more than 20 years with remarkable efficacy. HQC has been experimentally shown to exert anti-arthritic effects via the Wnt signaling pathway. AIM OF THE STUDY: This study used clinical data mining, network analysis, and in vitro and in vivo tests to investigate the anti-arthritic and possible anti-inflammatory mechanism of HQC...

Heat-shocked cells prioritize the translation of heat shock (HS) mRNAs, but the underlying mechanism is unclear. We report that HS in budding yeast induces the disassembly of the eIF4F complex, where eIF4G and eIF4E assemble into translationally arrested mRNA ribonucleoprotein particles (mRNPs) and HS granules (HSGs), whereas eIF4A promotes HS translation. Using in vitro reconstitution biochemistry, we show that a conformational rearrangement of the thermo-sensing eIF4A-binding domain of eIF4G dissociates eIF4A and promotes the assembly with mRNA into HS-mRNPs, which recruit additional translation factors, including Pab1p and eIF4E, to form multi-component condensates...

Regulation of mRNA translation by eukaryotic initiation factors (eIFs) is crucial for cell survival. In humans, eIF3 stimulates translation of the JUN mRNA which encodes the transcription factor JUN, an oncogenic transcription factor involved in cell cycle progression, apoptosis, and cell proliferation. Previous studies revealed that eIF3 activates translation of the JUN mRNA by interacting with a stem loop in the 5' untranslated region (5' UTR) and with the 5' -7-methylguanosine cap structure. In addition to its interaction site with eIF3, the JUN 5' UTR is nearly one kilobase in length, and has a high degree of secondary structure, high GC content, and an upstream start codon (uAUG)...

Non-small cell lung cancer (NSCLC) is the most common type of lung tumor; however, we lack effective early detection indicators and therapeutic targets. Eukaryotic translation initiation factor 4 gamma 1 (EIF4G1) is vital to initiate protein synthesis, acting as a scaffolding protein for the eukaryotic protein translation initiation factor complex, EIF4F, which regulates protein synthesis together with EIF4A, EIF4E, and other translation initiation factors. However, EIF4G1's function in NSCLC cancer is unclear...

G3BP1 is an RNA binding protein that condenses untranslating messenger RNAs into stress granules (SGs). G3BP1 is inactivated by multiple viruses and is thought to antagonize viral replication by SG-enhanced antiviral signaling. Here, we show that neither G3BP1 nor SGs generally alter the activation of innate immune pathways. Instead, we show that the RNAs encoded by West Nile virus, Zika virus, and severe acute respiratory syndrome coronavirus 2 are prone to G3BP1-dependent RNA condensation, which is enhanced by limiting translation initiation and correlates with the disruption of viral replication organelles and viral RNA replication...

Eukaryotic translation initiation involves recruitment of the 43S pre-initiation complex to the 5' end of mRNA by the cap-binding complex eIF4F, forming the 48S translation initiation complex (48S), which then scans along the mRNA until the start codon is recognized. We have previously shown that eIF4F binds near the mRNA exit channel of the 43S, leaving open the question of how mRNA secondary structure is removed as it enters the mRNA channel on the other side of the 40S subunit. Here we report the structure of a human 48S that shows that, in addition to the eIF4A that is part of eIF4F, there is a second eIF4A helicase bound at the mRNA entry site, which could unwind RNA secondary structures as they enter the 48S...

In this study, we aimed to address the current limitations of therapies for macro-metastatic triple-negative breast cancer (TNBC) and provide a therapeutic lead that overcomes the high degree of heterogeneity associated with this disease. Specifically, we focused on well-documented but clinically underexploited cancer-fueling perturbations in mRNA translation as a potential therapeutic vulnerability. We therefore developed an orally bioavailable rocaglate-based molecule, MG-002, which hinders ribosome recruitment and scanning via unscheduled and non-productive RNA clamping by the eukaryotic translation initiation factor (eIF) 4A RNA helicase...

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Non-canonical translation mechanisms have been described for many viral RNAs. In the case of several plant viruses, their protein synthesis is controlled by RNA elements in their genomic 3'-ends that are able to enhance cap-independent translation (3'-CITE). The proposed general mechanism of 3'-CITEs includes their binding to eukaryotic translation initiation factors (eIFs) that reach the 5'-end and AUG start codon through 5'-3'-UTR-interactions. It was previously shown that cucurbit aphid-borne yellows virus (CABYV) has a 3'-CITE, which varies in sequence and structure depending on the phylogenetic group to which the isolate belongs, possibly as a result of adaptation to the different geographical regions...

Inclusion bodies (IBs) of respiratory syncytial virus (RSV) are formed by liquid-liquid phase separation (LLPS) and contain internal structures termed "IB-associated granules" (IBAGs), where anti-termination factor M2-1 and viral mRNAs are concentrated. However, the mechanism of IBAG formation and the physiological function of IBAGs are unclear. Here, we found that the internal structures of RSV IBs are actual M2-1-free viral messenger ribonucleoprotein (mRNP) condensates formed by secondary LLPS. Mechanistically, the RSV nucleoprotein (N) and M2-1 interact with and recruit PABP to IBs, promoting PABP to bind viral mRNAs transcribed in IBs by RNA-recognition motif and drive secondary phase separation...

Initiation of translation is the first of the three obligatory steps required for protein synthesis and is carried out by a large number of protein factors called initiation factors in conjunction with ribosomes. One of the key conserved protein factors in eukaryotes that plays a role in this process is eIF4A, which has three homologues in humans with eIF4A1 being the primary factor playing a role in translation initiation. eIF4As are members of the family of DEAD-box helicases that carry out different biological functions...

Regulation of mRNA translation by eukaryotic initiation factors (eIFs) is crucial for cell survival. In humans, eIF3 stimulates translation of the JUN mRNA which encodes the transcription factor JUN, an oncogenic transcription factor involved in cell cycle progression, apoptosis, and cell proliferation. Previous studies revealed that eIF3 activates translation of the JUN mRNA by interacting with a stem loop in the 5' untranslated region (5' UTR) and with the 5' -7-methylguanosine cap structure. In addition to its interaction site with eIF3, the JUN 5' UTR is nearly one kilobase in length, and has a high degree of secondary structure, high GC content, and an upstream start codon (uAUG)...

The nuanced heterogeneity and specialized functions of translation machinery are increasingly recognized as crucial for precise translational regulation. Here, high-throughput ribosomal profiling (ribo-seq) is used to analyze the specialized roles of eukaryotic initiation factors (eIFs) in the budding yeast. By examining changes in ribosomal distribution across the genome resulting from knockouts of eIF4A, eIF4B, eIF4G1, CAF20, or EAP1, or knockdowns of eIF1, eIF1A, eIF4E, or PAB1, two distinct initiation-factor groups, the "looping" and "scanning" groups are discerned, based on similarities in the ribosomal landscapes their perturbation induced...

We have developed a deep sequencing-based approach, Rec-Seq, that allows simultaneous monitoring of ribosomal 48S pre-initiation complex (PIC) formation on every mRNA in the translatome in an in vitro reconstituted system. Rec-Seq isolates key early steps in translation initiation in the absence of all other cellular components and processes. Using this approach we show that the DEAD-box ATPase Ded1 promotes 48S PIC formation on the start codons of >1000 native mRNAs, most of which have long, structured 5'-untranslated regions (5'UTRs)...

Protein synthesis is frequently dysregulated in cancer and selective inhibition of mRNA translation represents an attractive cancer therapy. Here, we show that therapeutically targeting the RNA helicase eIF4A with zotatifin, the first-in-class eIF4A inhibitor, exerts pleiotropic effects on both tumor cells and the tumor immune microenvironment in a diverse cohort of syngeneic triple-negative breast cancer (TNBC) mouse models. Zotatifin not only suppresses tumor cell proliferation but also directly repolarizes macrophages toward an M1-like phenotype and inhibits neutrophil infiltration, which sensitizes tumors to immune checkpoint blockade...

Eukaryotic translation initiation factor 4E (eIF4E) has been known to play a critical role in the regulation of gene expression and essential cellular processes such as proliferation, apoptosis and differentiation. We explored herein its role in the pathophysiology of psoriasis. The inhibition of eIF4E by siRNA or briciclib, an eIF4E small molecule inhibitor, downregulated the expression of eIF4E itself and its two complex partners eIF4A and G, as well as other eIFs (e.g. eIF1A, eIF2α, eIF3A, eIF3B, eIF5 and eIF6)...

UNLABELLED: Protein synthesis is frequently dysregulated in cancer and selective inhibition of mRNA translation represents an attractive cancer therapy. Here, we show that therapeutically targeting the RNA helicase eIF4A by Zotatifin, the first-in-class eIF4A inhibitor, exerts pleiotropic effects on both tumor cells and the tumor immune microenvironment in a diverse cohort of syngeneic triple-negative breast cancer (TNBC) mouse models. Zotatifin not only suppresses tumor cell proliferation but also directly repolarizes macrophages towards an M1-like phenotype and inhibits neutrophil infiltration, which sensitizes tumors to immune checkpoint blockade...