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Updated: May 17, 2026

Identification of Alternative Splicing and Polyadenylation in RNA-seq Data
Published on: June 24, 2021
Transcriptome-wide analyses of CstF64-RNA interactions in global regulation of mRNA alternative polyadenylation
Chengguo Yao1, Jacob Biesinger, Ji Wan
1Department of Microbiology and Molecular Genetics, School of Medicine, University of California, Irvine, CA 92697, USA.
Abstract:
Cleavage stimulation factor 64 kDa (CstF64) is an essential pre-mRNA 3' processing factor and an important regulator of alternative polyadenylation (APA). Here we characterized CstF64-RNA interactions in vivo at the transcriptome level and investigated the role of CstF64 in global APA regulation through individual nucleotide resolution UV crosslinking and immunoprecipitation sequencing and direct RNA sequencing analyses. We observed highly specific CstF64-RNA interactions at poly(A) sites (PASs), and we provide evidence that such interactions are widely variable in affinity and may be differentially required for PAS recognition. Depletion of CstF64 by RNAi has a relatively small effect on the global APA profile, but codepletion of the CstF64 paralog CstF64τ leads to greater APA changes, most of which are characterized by the increased relative use of distal PASs. Finally, we found that CstF64 binds to thousands of dormant intronic PASs that are suppressed, at least in part, by U1 small nuclear ribonucleoproteins. Taken together, our findings provide insight into the mechanisms of PAS recognition and identify CstF64 as an important global regulator of APA.
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