Splicing factor U2AF2 promotes inclusion of exons with poor nucleosome phasing

Spliceosome recognition of pre-mRNA junctions requires binding of splicing factors to the 3' splice site. Most splicing occurs co-transcriptionally, but how splicing factors bind expressed gene is not understood. We show that the SF3B1, U2AF1, and U2AF2 splicing factors bind active promoters of both intron-containing and intronless genes. RNase A treatment reduces binding at promoters, implicating general interactions with nascent transcripts in promoter localization. Strikingly, U2AF2 also accumulates on the bodies of some genes and is not eliminated by RNase treatment. Neither RNA polymerase II nor the active gene body chromatin mark H3K36me3 is required to maintain U2AF2 on chromatin. We found that most exons in U2AF2-bound have phased nucleosomes at their exon-intron boundaries. However, some exons are poorly phased, and these require U2AF2 for inclusion in mature mRNA. We propose that nucleosome phasing and U2AF2 binding ensure exon inclusion. We used Cleavage under targets and Release using nuclease (Cut-and-Run), a chromatin profiling strategy in which antibody-targeted controlled cleavage by micrococcal nuclease releases specific protein-DNA complexes into the supernatant for paired-end DNA sequencing