eIF4A2 drives repression of translation at initiation by Ccr4-Not through purine-rich motifs in the 5′UTR

Abstract

Background: Regulation of the mRNA life cycle is central to gene expression control and determination of cell fate. miRNAs represent a critical mRNA regulatory mechanism, but despite decades of research, their mode of action is still not fully understood. Results: Here, we show that eIF4A2 is a major effector of the repressive miRNA pathway functioning via the Ccr4-Not complex. We demonstrate that while DDX6 interacts with Ccr4-Not, its effects in the mechanism are not as pronounced. Through its interaction with the Ccr4-Not complex, eIF4A2 represses mRNAs at translation initiation. We show evidence that native eIF4A2 has similar RNA selectivity to chemically inhibited eIF4A1. eIF4A2 exerts its repressive effect by binding purine-rich motifs which are enriched in the 5′UTR of target mRNAs directly upstream of the AUG start codon. Conclusions: Our data support a model whereby purine motifs towards the 3′ end of the 5′UTR are associated with increased ribosome occupancy and possible uORF activation upon eIF4A2 binding.

General and Target-Specific DExD/H RNA Helicases in Eukaryotic Translation Initiation

“…EIF4A2 has been implicated in translational repression and microRNA (miRNA) regulation through its interactions with the Ccr4-Not complex [51,52]. This complex is recruited to mRNAs via PABP, miRNAs, and RNA-binding proteins (RBPs), among others, when an mRNA is targeted for deadenylation and decay [53][54][55][56].…”

“…Meijer et al [57] first demonstrated that eIF4A2 activity is required for miRNA-mediated mRNA destabilization. Then, the same lab reported that eIF4A2 may exert this function by becoming incorporated into the Ccr4-Not complex and inhibiting the CNOT7 deadenylation activity or repressing translation by binding to purine-rich sequences proximal to the AUG start codon [51,52]. These experiments implicating a repressive function for eIF4A2 are at odds with eIF4A2 being able to incorporate into the eIF4F complex, which plays a stimulatory role in translation [58].…”

So close, no matter how far: multiple paths connecting transcription to mRNA translation in eukaryotes

“…Second, CCR4-Not promotes the degradation of inefficiently translated mRNAs by monitoring decoding rates (Webster et al, 2018;Buschauer et al, 2020). Third, together with the eIF4A2 protein, it may participate in the miRNAmediated inhibition of translation initiation of certain transcripts possessing purine-rich sequences within their 5 0 UTR (Wilczynska et al, 2019). Thus, the CCR4-Not complex is an important regulator of multiple processes, and its co-transcriptional recruitment to the nascent transcripts may profoundly impact their translation, as shown for the yeast ribosomal proteins (Gupta et al, 2016).…”

The Regulatory Properties of the Ccr4–Not Complex

“…Initially it was shown that four yeast Not proteins were involved in the global repression of transcription initiation [11,155]. However, it now seems reasonable to suppose that the whole complex plays a role in the process [123,156]. The Ccr4-Not complex functionally and physically associates with TBP and TBP-associated components [152].…”