mRNA 50 terminal sequences drive 200-fold differences in expression through effects on synthesis, translation and decay

Abstract

mRNA regulatory sequences control gene expression at multiple levels including translation initiation and mRNA decay. The 50 terminal sequences of mRNAs have unique regulatory potential because of their proximity to key post-transcriptional regulators. Here we have systematically probed the function of 50 terminal sequences in gene expression in human cells. Using a library of reporter mRNAs initiating with all possible 7-mer sequences at their 50 ends, we find an unexpected impact on transcription that underlies 200-fold differences in mRNA expression. Library sequences that promote high levels of transcription mirrored those found in native mRNAs and define two basic classes with similarities to classic Initiator (Inr) and TCT core promoter motifs. By comparing transcription, translation and decay rates, we identify sequences that are optimized for both efficient transcription and growth-regulated translation and stability, including variants of terminal oligopyrimidine (TOP) motifs. We further show that 50 sequences of endogenous mRNAs are enriched for multifunctional TCT/TOP hybrid sequences. Together, our results reveal how 50 sequences define two general classes of mRNAs with distinct growth-responsive profiles of expression across synthesis, translation and decay.