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Background: Alternative polyadenylation (APA) affects most human genes and is recurrently dysregulated in all studied cancers. However, the mechanistic origins of this dysregulation are incompletely understood. Results: We describe an unbiased analysis of molecular regulators of poly(A) site selection across The Cancer Genome Atlas and identify that colorectal adenocarcinoma is an outlier relative to all other cancer subtypes. This distinction arises from the frequent presence of loss-of-function APCmutations in colorectal adenocarcinoma, which are strongly associated with long 3′ UTR expression relative to tumors lacking APCmutations. APCknockout similarly dysregulates APA in human colon organoids. By mining previously published APC eCLIP data, we show that APC preferentially binds G- and C-rich motifs just upstream of proximal poly(A) sites. Lastly, we find that reduced APCexpression is associated with APA dysregulation in tumor types lacking recurrent APCmutations. Conclusions: As APC has been previously identified as an RNA-binding protein that preferentially binds 3′ UTRs during mouse neurogenesis, our results suggest that APC promotes proximal poly(A) site use and that APC loss and altered expression contribute to pervasive APA dysregulation in cancers.

Most eukaryotes harbor two distinct pre-mRNA splicing machineries: the major spliceosome, which removes >99% of introns, and the minor spliceosome, which removes rare, evolutionarily conserved introns. Although hypothesized to serve important regulatory functions, physiologic roles of the minor spliceosome are not well understood. For example, the minor spliceosome component ZRSR2 is subject to recurrent, leukemia-associated mutations, yet functional connections among minor introns, hematopoiesis and cancers are unclear. Here, we identify that impaired minor intron excision via ZRSR2 loss enhances hematopoietic stem cell self-renewal. CRISPR screens mimicking nonsense-mediated decay of minor intron-containing mRNA species converged on LZTR1, a regulator of RAS-related GTPases. LZTR1minor intron retention was also discovered in the RASopathy Noonan syndrome, due to intronic mutations disrupting splicing and diverse solid tumors. These data uncover minor intron recognition as a regulator of hematopoiesis, noncoding mutations within minor introns as potential cancer drivers and links among ZRSR2mutations, LZTR1 regulation and leukemias.

Pineda and Bradley demonstrate that almost all human introns contain multiple branchpoints. Approximately three-quarters of constitutive introns exhibit tissue-specific branchpoint usage.