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

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. The translational impact of the compounds including Hipp, San, or DMDA-PatA was examined through ribosome profiling. RNA pulldown-Seq was conducted to explore the interaction between mRNAs and eIF4A1 in the presence of DMDA-PatA. RNA Bind-n-Seq was performed to elucidate the sequence preferences of RNA binding proteins in the presence of DMDA-PatA, iPr-DMDA-PatA, or RocA.