On the rules of engagement for microRNAs targeting protein coding regions

Abstract

MiRNAs post-transcriptionally repress gene expression by binding to mRNA 3'UTRs, but the extent to which they act through protein coding regions (CDS regions) is less well established. MiRNA interaction studies show a substantial proportion of binding occurs in CDS regions, however sequencing studies show much weaker effects on mRNA levels than from 3'UTR interactions, presumably due to competition from the translating ribosome. Consequently, most target prediction algorithms consider only 3'UTR interactions. However, the consequences of CDS interactions may have been underestimated, with the reporting of a novel mode of miRNA-CDS interaction requiring base pairing of the miRNA 3' end, but not the canonical seed site, leading to repression of translation with little effect on mRNA turnover. Using extensive reporter, western blotting and bioinformatic analyses, we confirm that miRNAs can indeed suppress genes through CDS-interaction in special circumstances. However, in contrast to that previously reported, we find repression requires extensive base-pairing, including of the canonical seed, but does not strictly require base pairing of the 3' miRNA terminus and is mediated through reducing mRNA levels. We conclude that suppression of endogenous genes can occur through miRNAs binding to CDS, but the requirement for extensive base-pairing likely limits the regulatory impacts to modest effects on a small subset of targets.