Head-on and co-directional RNA polymerase collisions orchestrate bidirectional transcription termination

Genomic DNA is a crowded track where translocating motor proteins frequently collide. It remains unclear whether these collisions, generally thought to occur inadvertently, carry any physiological function. In this work, we developed a single-molecule assay to directly visualize the trafficking of individual E. coli RNA polymerases (RNAPs) on DNA. This assay enabled us to test the hypothesis that RNAP collisions drive bidirectional transcription termination of convergent gene pairs. We showed that the head-on collision between two converging RNAPs is necessary to prevent transcriptional readthrough, but insufficient to release the collided RNAPs from the DNA. Remarkably, co-directional collision from a trailing RNAP into the head-on collided complex dramatically increases the termination efficiency. Furthermore, a stem-loop structure formed in the nascent RNA is required for collisions to occur at well-defined positions between gene boundaries. These findings, corroborated by transcriptomic data, establish programmed RNAP collisions as an effective strategy to achieve precise gene expression and imply a broader role of genomic conflicts in cell physiology. ### Competing Interest Statement The authors have declared no competing interest.