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3′ end processing is required for the maturation of all eukaryotic RNAs. Current model suggests that canonical mRNA 3′ processing is carried out exclusively within a protein complex termed mRNA 3′ processing complex. In a recent study, by using RNA-biotin based pull-down assay and high-throughput sequencing, we reported that a subset of small nucleolar RNAs (snoRNAs) were physically associated with this macromolecular machinery. Through detailed characterization of one of these snoRNAs, SNORD50A, we revealed that non-coding RNA, such as snoRNA, may play a regulatory role in mRNA 3′ processing. Our results provided novel insight into both the regulatory mechanism of mRNA 3′ processing and the non-canonical functions of snoRNAs.

mRNA 3′ end processing is an essential step in gene expression. It is well established that canonical eukaryotic pre-mRNA 3′ processing is carried out within a macromolecular machinery consisting of dozens of trans-acting proteins. However, it is unknown whether RNAs play any role in this process. Unexpectedly, we found that a subset of small nucleolar RNAs (snoRNAs) are associated with the mammalian mRNA 3′ processing complex. These snoRNAs primarily interact with Fip1, a component of cleavage and polyadenylation specificity factor (CPSF). We have functionally characterized one of these snoRNAs and our results demonstrated that the U/A-rich SNORD50A inhibits mRNA 3′ processing by blocking the Fip1-poly(A) site (PAS) interaction. Consistently, SNORD50A depletion altered the Fip1–RNA interaction landscape and changed the alternative polyadenylation (APA) profiles and/or transcript levels of a subset of genes. Taken together, our data revealed a novel function for snoRNAs and provided the first evidence that non-coding RNAs may play an important role in regulating mRNA 3′ processing.

It is increasingly appreciated that U1 snRNP transcriptomically suppresses the usage of intronic polyadenylation site (PAS) of mRNAs, an outstanding question is why frequently used PASs are not suppressed. Here we found that U1 snRNP could be transiently associated with sequences upstream of actionable PASs in human cells, and RNA-RNA interaction might contribute to the association. By focusing on individual PAS, we showed that the stable assembly of U1 snRNP near PAS might be generally required for U1 inhibition of mRNA 3' processing. Therefore, actionable PASs that often lack optimal U1 snRNP docking site nearby is free from U1 inhibitory effect. Consistently, natural 5' splicing site (5'-SS) is moderately enriched ~250 nt upstream of intronic PASs whose usage is sensitive to functional knockdown of U1 snRNA. Collectively, our results provided an insight into how U1 snRNP selectively inhibits the usage of PASs in a cellular context, and supported a prevailing model that U1 snRNP scans pre-mRNA through RNA-RNA interaction to find a stable interaction site to exercise its function in pre-mRNA processing, including repressing the usage of cryptic PASs.