Full-length RNA profiling reveals pervasive bidirectional transcription terminators in bacteria

The ability to determine full-length nucleotide composition of individual RNA molecules is essential for understanding the architecture and function of a transcriptome. However, experimental approaches capable of capturing the sequences of both 5’ and 3’ termini of the same transcript remain scarce. Here we present SEnd-seq—a high-throughput and unbiased method that simultaneously maps transcription start and termination sites with single-nucleotide resolution. Using this method, we obtain a comprehensive view of the Escherichia coli transcriptome, which displays an unexpected level of complexity. SEnd-seq significantly expands the catalog of transcription start sites and termination sites, defines unique transcription units, and detects prevalent antisense RNA. Strikingly, our results unveil widespread overlapping bidirectional terminators located between opposing gene pairs. We further show that convergent transcription is a major contributor to highly efficient bidirectional termination both in vitro and in vivo. This finding highlights an underappreciated role of RNA polymerase conflicts in shaping transcript boundaries and suggests an evolutionary strategy for modulating transcriptional output by arranging gene orientation.