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Pateamine A mediates RNA sequence-selective translation repression by anchoring eIF4A and DDX3 to GNG motifs

ABSTRACT: Small molecule compounds that inducespark the mRNA-selective translation repression, are attracting interesthave arisen due to their potential for expansion ofin druggable space expansion. However, only limited examples have been reported to datewere found. Here we showed that pateamine A (PatA) represses translation in an mRNA-selective manner by clamping eIF4A, a DEAD-box RNA-binding protein, on GNG motifs. Through systematic comparison of multiple eIF4A inhibitors by ribosome profiling, we found that PatA has a unique mRNA selectivity in translation repression. Unbiased Bind-n-Seq revealed that PatA provides a sequence preference toward GNG motifs to eIF4A in an ATP-independent manner. This unusual RNA binding sterically hindered scanning by 40S ribosomes. In silico simulation, quantum chemical calculation, and subsequent development of an inactive PatA derivative concluded that the charge of the secondary amine on the trienyl arm determines G selectivity. Moreover, we found that DDX3, another DEAD-box protein, was an alternative target of PatA, phenocopying the effect on eIF4A. Our results provide an example of sequence-selective anchor of RNA-binding proteins and mRNA-selective inhibition of protein synthesis by a small molecule compound.

ORGANISM(S): synthetic construct Homo sapiens

PROVIDER: GSE243312 | GEO | 2024/07/08

REPOSITORIES: GEO

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Project description:DEAD-box RNA helicases eIF4A and Ded1 are believed to promote translation initiation by resolving mRNA secondary structures that impede ribosome attachment at the mRNA 5’ end or subsequent scanning of the 5’UTR, but whether they perform distinct functions or act redundantly in vivo is poorly understood. We compared the effects of mutations in Ded1 or eIF4A on global translational efficiencies (TEs) in yeast by ribosome footprint profiling. Despite similar reductions in bulk translation, inactivation of a cold-sensitive Ded1 mutant substantially reduced the TEs of >600 mRNAs, whereas inactivation of a temperature-sensitive eIF4A mutant yielded <40 similarly impaired mRNAs. The broader requirement for Ded1 did not reflect more pervasive secondary structures at low temperature, as inactivation of temperature-sensitive and cold-sensitive ded1 mutants gave highly correlated results. Interestingly, Ded1-dependent mRNAs exhibit greater than average 5’UTR length and propensity for secondary structure, implicating Ded1 in scanning though structured 5' UTRs. Reporter assays confirmed that cap- distal stem-loop insertions increase dependence on Ded1 but not eIF4A for efficient translation. While only a small fraction of mRNAs is strongly dependent on eIF4A, this dependence is significantly correlated with requirements for Ded1 and 5’UTR features characteristic of Ded1- dependent mRNAs. Our findings suggest that Ded1 is critically required to promote scanning through secondary structures within 5’UTRs; and while eIF4A cooperates with Ded1 in this function, it also promotes a step of initiation common to virtually all yeast mRNAs. We compared the effects of mutations in Ded1 or eIF4A on global translational efficiencies (TEs) in yeast by ribosome footprint profiling.The study includes 32 samples, comprised of 16 mRNA-Seq samples and 16 ribosome footprint profiling samples, derived from biological replicates of 3 mutant strains, ded1-cs, ded1-ts and tif1-ts, and the corresponding wild-type strains. The tif1-ts mutant and its wild-type counterpart were analyzed at 30°C and 37°C.

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Pateamine A mediates RNA sequence-selective translation repression by anchoring eIF4A and DDX3 to GNG motifs

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