Widespread Influence of 3'-end Structures on Mammalian mRNA Processing and Stability [4sU-2P-seq]

Understanding the physiological relevance of structures in mammalian mRNAs remains elusive, especially considering the global unfolding of mRNA structures in eukaryotic organisms recently examined, as well as the decade-long observation that mRNAs generally seem no more likely than random sequences to be stably folded. Here we show that RNA secondary structures, mostly weak and close-to-random, facilitate the 3'-end processing of thousands of human mRNAs by juxtaposing poly(A) signals (PASs) and cleavage sites that are otherwise too far apart. Folding of these 3'-end structures also enhances mRNA stability. Global structure probing shows that 3'-end regions are indeed folded in cells despite substantial unfolding of PAS-upstream regions. Analyses of thousands of ectopically expressed variants prove that folding both enhances processing and increases stability. Mutagenesis of a genomic locus further implicates structure-controlled processing in regulating neighboring gene expression. These results reveal widespread roles for RNA structure in mammalian mRNA biogenesis and metabolism. Overall design: Three samples: steady-state, nascent RNA labeled by 4sU for 30 min, and labeled for 60 min