EIF4A1 enhances LARP1-mediated translational repression during mTORC1 inhibition
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PROVIDER: PRJNA763797 | ENA |
REPOSITORIES: ENA
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Project description:Eukaryotic translation initiation factor (eIF) 4A — a DEAD-box RNA-binding protein — plays an essential role in translation initiation. Recent reports have suggested helicase-dependent and helicase-independent functions for eIF4A, but the multifaceted roles of eIF4A have not been fully explored. Here, we show that eIF4A1 enhances translational repression during the inhibition of mechanistic target of rapamycin complex 1 (mTORC1), an essential kinase complex controlling cell proliferation. RNA pulldown followed by sequencing revealed that eIF4A1 preferentially binds to mRNAs containing terminal oligopyrimidine (TOP) motifs (TOP mRNAs), whose translation is rapidly repressed upon mTORC1 inhibition. This selective interaction depends on a La-related RNA-binding protein, LARP1. Ribosome profiling revealed that deletion of EIF4A1 attenuated the translational repression of TOP mRNAs upon mTORC1 inactivation. Moreover, eIF4A1 increases the affinity between TOP mRNAs and LARP1 and thus ensures stronger translational repression upon mTORC1 inhibition. Our data show the multimodality of eIF4A1 in modulating protein synthesis through an inhibitory binding partner and provide a unique example of the repressive role of a universal translational activator.
2024-02-13 | GSE184247 | GEO
Project description:The RNA biding protein, LARP1, has been proposed to function downstream of mTORC1 to positively regulate the translation of 5M-bM-^@M-^YTOP mRNAs such as ribosome protein (RP) mRNAs. However, its regulatory roles in mTORC1-mediated translation remain unclear. PAR-CLIP of LARP1 revealed its direct and dynamic interactions with RP mRNAs through pyrimidine-enriched sequences in the 5M-bM-^@M-^YUTR of RP mRNAs when mTOR activity is inhibited. Importantly, this LARP1 is a direct substrate of mTORC1 and S6K1/Akt, and phosphorylated LARP1 scaffolds mTORC1 on translation-competent mRNAs to facilitate 4EBP1 and S6K1 phosphorylation. Ablation of LARP1 causes multiple defects in the processes of translation including abnormal eIF4G1 interaction with RP mRNAs and inefficient RP mRNA elongation thereby reducing ribosome biogenesis and cell proliferation. These observations illustrate that LARP1 functions both an effector and a regulator for local mTORC1 activity, and acts as a molecular switch for ribosome biogenesis by sensing growth factor/nutrient signaling. LARP1-bound RNA regions were sequenced from HEK293T cells under growing or mTOR-inactive conditions. In parallel, mRNA abundance was quantified, in biological duplicate, from HEK293T cells under the same conditions.
2014-07-31 | E-GEOD-59599 | biostudies-arrayexpress
Project description:The RNA biding protein, LARP1, has been proposed to function downstream of mTORC1 to positively regulate the translation of 5’TOP mRNAs such as ribosome protein (RP) mRNAs. However, its regulatory roles in mTORC1-mediated translation remain unclear. PAR-CLIP of LARP1 revealed its direct and dynamic interactions with RP mRNAs through pyrimidine-enriched sequences in the 5’UTR of RP mRNAs when mTOR activity is inhibited. Importantly, this LARP1 is a direct substrate of mTORC1 and S6K1/Akt, and phosphorylated LARP1 scaffolds mTORC1 on translation-competent mRNAs to facilitate 4EBP1 and S6K1 phosphorylation. Ablation of LARP1 causes multiple defects in the processes of translation including abnormal eIF4G1 interaction with RP mRNAs and inefficient RP mRNA elongation thereby reducing ribosome biogenesis and cell proliferation. These observations illustrate that LARP1 functions both an effector and a regulator for local mTORC1 activity, and acts as a molecular switch for ribosome biogenesis by sensing growth factor/nutrient signaling.
2014-07-31 | GSE59599 | GEO
Project description:LARP1 has been proposed to control the translation of TOP mRNAs downstrteam of mTORC1. Here we used ribosome profiling to analyze transcriptome-wide changes in translation following mTOR inhibition in wild-type HEK-293T cells and cells where LARP1 (sgLARP1) or LARP1 and its homologue LARP1B (sgLARP1/1B) have been deleted using CRISPR/Cas9.
2020-02-12 | GSE132703 | GEO
Project description:La-related protein 1 (LARP1) has been identified as a key translational inhibitor of terminal oligopyrimidine tract (TOP) mRNAs downstream of the nutrient sensing protein kinase complex, mTORC1. LARP1 exerts this inhibitory effect on TOP mRNA translation by binding to the mRNA cap and the adjacent 5’TOP motif, resulting in the displacement of the eIF4E complex from TOP mRNAs. In the present study, we identify a second nutrient sensing kinase GCN2 that converges on LARP1 to control TOP mRNA translation. GCN2 inhibits TOP mRNA translation via ATF4-dependent transcriptional induction of LARP1 mRNAs and GCN1-mediated recruitment of LARP1 to stalled ribosomes. We performed ATF4 ChIP-seq experiments in both WT and GCN2 KO MEFs with or without leucine deprivation.
2022-08-31 | GSE166590 | GEO
Project description:mTORC1 controls a dynamic program of TOP mRNA translation via LARP1
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Project description:Hippuristanol (Hipp) is a natural product that selectively inhibits protein synthesis by targeting eukaryotic initiation factor (eIF) 4A, a DEAD-box RNA helicase required for ribosome recruitmentt o mRNA templates. Using a CRISPR/Cas9-based variomics screen, we identify functional eIF4A1 Hipp-resistant alleles, which in turn allow us to link the translation-inhibitory and cytotoxic properties of Hipp to eIF4A1 target-engagement. Genome-wide translational profiling in the absence or presence of Hipp (~EC50) were undertaken and our validation studies provided insight into structure-activity relationships of eIF4A-dependent mRNAs.
2020-08-05 | GSE151687 | GEO
Project description:We screened the translational targets of eIF4A1 in DU145 cells using the Native RNA immunoprecipitation (RIP) assay with RNA-seq (RIP-seq). The eIF4A1-binding peaks and RNA fractions were normally distributed around the ATG translation start site. A total of 197 coding genes with eFI4A1-binding peaks in mRNAs, including 5' UTR, exon, and 3' UTR regions. were identified in the eIF4A1-RIP sample. The most enriched eFI4A1-binding motifs (MAGGTA, CCASCYC, and GARGA) were identified by aligning the sequences of all eFI4A1-binding RNA fractions to a reference genome.
2022-06-29 | GSE161709 | GEO
Project description:Oncogenic translational programmes are an emerging hallmark of cancer and often driven by dysregulation of signaling pathways including KRAS and mTORC that converge on the eukaryotic translation initiation (eIF) 4F complex. Altered eIF4F activity promotes translation of oncogene mRNAs that typically contain highly structured 5’UTRs rendering their translation strongly dependent on RNA unwinding by DEAD-box helicase eIF4A1 subunit of the eIF4F complex. In addition, eIF4A1 separately functions to load mRNA into the 43S pre-initiation complex (PIC), an essential step for the translation of cellular mRNA. While eIF4A1-dependent mRNAs have been widely investigated, it is still unclear if highly structured mRNAs recruit and activate eIF4A1 unwinding specifically. Here, we uncover that unwinding by eIF4A1 is activated in an RNA sequence-dependent manner in cells. Our data demonstrate that eIF4A1-dependent mRNAs contain specific RNA sequences, particularly enriched for polypurine-motifs, in their 5’UTR which recruit and specifically stimulate unwinding of local repressive RNA structure by eIF4A1 in an RNA sequence-dependent manner to facilitate translation. Mechanistically, we show that polypurine-rich sequences trigger the formation of RNA sequence-specific multimeric eIF4A1-complexes, assembled of catalytically distinct eIF4A1 subunits, the joint activity of which enhances RNA unwinding activity. Together with our structural data, we describe a model in which conformational changes within eIF4A1 and the RNA through the process of eIF4A1 multimerisation, lead to an optimal interaction of eIF4A1-unwinding subunits with the structured RNA region which enhances unwinding. Hence, we conclude that RNA sequences in addition to protein cofactors contribute to the regulation of cellular eIF4A1 function and promotion of translation of eIF4A1-unwinding dependent mRNAs.
2023-01-17 | PXD034343 | Pride
Project description:EIF4A1 and cofactors EIF4B and EIF4H have been well characterised in cancers, including B cell malignancies, for their ability to promote the translation of oncogenes with structured 5’ untranslated regions but very little is known of their roles in non-malignant cells. Using mouse models to delete Eif4a1, Eif4b or Eif4h in B cells we show that EIF4A1, but not EIF4B or EIF4H, is essential for B cell development and the germinal centre response. Following activation, EIF4A1 facilitates an increased rate of protein synthesis, MYC expression and expression of cell cycle regulators. However, EIF4A1-deficient cells remain viable whereas Hippuristanol treatment induces cell death.
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