DSIF factor Spt5 coordinates transcription, maturation and exoribonucleolysis of RNA polymerase II transcripts
RNA polymerase II (Pol II) termination is a crucial step in the transcriptional cycle, ensuring the recycling of Pol II and preventing interference with neighboring gene transcription. Xrn2 is an enzyme with 5'-3'- exonuclease activity, responsible for degrading RNA that is still being transcribed by Pol II after cleavage at the poly(A) site, ultimately leading to Pol II termination. We have confirmed that Xrn2's catalytic activity is essential for timely transcription termination. Furthermore, we have discovered a link between Xrn2 and the conserved, multifunctional transcription factor Spt5. Importantly, our research demonstrates that Xrn2 interacts with Pol II/Spt5 complexes. Additionally, when Spt5 is depleted, it not only impairs global transcription but also results in defective transcription termination. On the other hand, absence of Spt5 leads to the derepression of non-coding transcription and premature termination/attenuation at the 5'-end of coding genes. We propose that Spt5 acts as a "licensing" factor, ensuring that Pol II complexes are properly assembled for efficient transcription and co-transcriptional pre-mRNA processing. Overall design: Nascent transcriptome (TT-seq) was used to study how Spt5 and Xrn2 impact transcription. Spt5 was depleted using auxin degron and compared to DMSO-treated strain. Three Xrn2 degron strains were created: one without Xrn2 rescue, one with WT rescue, and one with MUT rescue (containing a catalytic D235A mutation). The rescue Xrn2 (or its mutant) under the native promoter was placed in the ura4 locus. The experiments were performed twice and spike-in normalized.