Coupling of Co-transcriptional Splicing and 3’ End Pol II Pausing During Termination in Arabidopsis
Published:09 Oct.2023    Source:Genome Biology
In Arabidopsis, RNA Polymerase II (Pol II) often pauses within a few hundred base pairs downstream of the polyadenylation site, reflecting efficient transcriptional termination, but how such pausing is regulated remains largely elusive.
Here, we analyze Pol II dynamics at 3 ends by combining comprehensive experiments with mathematical modelling. We generate high-resolution serine 2 phosphorylated (Ser2P) Pol II positioning data specifically enriched at 3 ends and define a 3 end pause index (3’ PI). The position but not the extent of the 3 end pause correlates with the termination window size. The 3’ PI is not decreased but even mildly increased in the termination deficient mutant xrn3, indicating 3 end pause is a regulatory step early during the termination and before XRN3-mediated RNA decay that releases Pol II. Unexpectedly, 3’ PI is closely associated with gene exon numbers and co-transcriptional splicing efficiency. Multiple exons genes often display stronger 3 end pauses and more efficient on-chromatin splicing than genes with fewer exons. Chemical inhibition of splicing strongly reduces the 3’ PI and disrupts its correlation with exon numbers but does not globally impact 3 end readthrough levels.

These results are further confirmed by fitting Pol II positioning data with a mathematical model, which enables the estimation of parameters that define Pol II dynamics. Our work highlights that the number of exons via co-transcriptional splicing is a major determinant of Pol II pausing levels at the 3’ end of genes in plants.