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

Genomic DNA is a crowded track where motor proteins frequently collide. It remains underexplored whether these collisions carry physiological function. In this work, we develop a single-molecule assay to visualize the trafficking of individual E. coli RNA polymerases (RNAPs) on DNA. Based on transcriptomic data, we hypoth-esize that RNAP collisions drive bidirectional transcription termination of convergent gene pairs. Single -molecule results show that the head-on collision between two converging RNAPs is necessary to prevent transcriptional readthrough but insufficient to release the RNAPs from the DNA. Remarkably, co-directional collision of a trailing RNAP into the head-on collided complex dramatically increases the termination effi-ciency. Furthermore, stem-loop structures formed in the nascent RNA are required for collisions to occur at well-defined positions between convergent genes. These findings suggest that physical collisions be-tween RNAPs furnish a mechanism for transcription termination and that programmed genomic conflicts can be exploited to co-regulate the expression of multiple genes.