Overlapping and distinctfunctions of SPT6, PNUTS and PCF11 in regulating transcription termination [ChIP-Seq]

The histone chaperone and transcription elongation factor SPT6, integral to RNA Polymerase II (RNAPII) activity, also plays a crucial role in regulating transcription termination. In an attempt to identify the pathways employed by SPT6 in this regulation, we found that while SPT6 and its closest partner IWS1 interact and co-localize with RNAPII, their functions diverge significantly at gene termination sites. Our data revealed that SPT6 depletion, unlike IWS1, results in extensive readthrough transcription, indicating that SPT6 independently regulates transcription termination. Further analysis identified that the cleavage and polyadenylation factor PCF11 and the phosphatase regulatory protein PNUTS collaborate with SPT6 in this process. These findings suggest that SPT6 may facilitate transcription termination by recruiting PNUTS and PCF11 to RNAPII. Additionally, SPT6 and PNUTS jointly restrict promoter upstream transcripts (PROMPTs), whereas PCF11 presence is essential for their activation in the absence of SPT6. This study elucidates the distinct and overlapping functions of SPT6, PCF11, and PNUTS in transcription termination, highlighting the pivotal role of SPT6 in ensuring proper transcription termination at the 5’ and 3’ ends of genes. Chromatin Immunoprecipitation sequencing (ChIP-seq) for SPT6, IWS1, PNUTS and RNAPII (siCT versus siPNUTS conditions) in HeLa cells