Functional coupling between ribosomal RNA transcription and processing guided by stable transcription factor binding

Coordinating ribosomal RNA (rRNA) transcription, folding and processing is essential for bacterial ribosome assembly. Yet, the molecular mechanisms underlying this coordination remain poorly understood. Particularly, whether the rRNA transcription antitermination complex (rrnTAC: NusA, NusG, NusB, NusE, S4, SuhB) orchestrates this coordination remains unclear. Here, we develop a suite of multi-color single-molecule fluorescence microscopy assays to simultaneously visualize, in real-time, rrnTAC assembly dynamics and their effect on transcription and co-transcriptional processing by RNase III. We find that transient ([~]1 s) interactions of general transcription factors NusA and NusG with RNAP, and of NusB/E with the rRNA-specific boxBAC element, are stabilized to minutes-long residence by final SuhB recruitment. Stable rrnTAC assembly is required to reduce RNAP pausing and to boost co-transcriptional rRNA processing. Collectively, our results reveal how transcription factor binding dynamics relate to function: transient for messenger RNA transcription and stable for rRNA transcription and processing.