Gene

Cell biology

Linguistics

Philosophy

Biology

Messenger RNA

RNA polymerase II

Cleavage (geology)

Intron

Transcription (linguistics)

Primary transcript

Promoter

Anemia

Paleontology

Exonic splicing enhancer

Genetics

Group II intron

Post-transcriptional modification

Erythropoiesis

Splicing factor

RNA splicing

Alternative splicing

Internal medicine

Precursor mRNA

Fracture (geology)

Medicine

Exon

Gene expression

Molecular Cell

Pre-mRNA processing steps are tightly coordinated with transcription in many organisms. To determine how co-transcriptional splicing is integrated with transcription elongation and 3' end formation in mammalian cells, we performed long-read sequencing of individual nascent RNAs and precision run-on sequencing (PRO-seq) during mouse erythropoiesis. Splicing was not accompanied by transcriptional pausing and was detected when RNA polymerase II (Pol II) was within 75-300 nucleotides of 3' splice sites (3'SSs), often during transcription of the downstream exon. Interestingly, several hundred introns displayed abundant splicing intermediates, suggesting that splicing delays can take place between the two catalytic steps. Overall, splicing efficiencies were correlated among introns within the same transcript, and intron retention was associated with inefficient 3' end cleavage. Remarkably, a thalassemia patient-derived mutation introducing a cryptic 3'SS improved both splicing and 3' end cleavage of individual β-globin transcripts, demonstrating functional coupling between the two co-transcriptional processes as a determinant of productive gene output.

Co-transcriptional splicing regulates 3′ end cleavage during mammalian erythropoiesis