A general mechanism for transcription bubble nucleation in bacteria

Bacterial transcription initiation requires s factors for nucleation of the transcription bubble. The canonical housekeeping s factor, sigma(70), nucleates DNA melting via recognition of conserved bases of the promoter -10 motif, which are unstacked and captured in pockets of sigma(70). By contrast, the mechanism of transcription bubble nucleation and formation during the unrelated sigma(N)-mediated transcription initiation is poorly understood. Herein, we combine structural and biochemical approaches to establish that sigma(N), like sigma(70), captures a flipped, unstacked base in a pocket formed between its N-terminal region I (RI) and extra-long helix features. Strikingly, RI inserts into the nascent bubble to stabilize the nucleated bubble prior to engagement of the obligate ATPase activator. Our data suggest a general paradigm of transcription initiation that requires s factors to nucleate an early melted intermediate prior to productive RNA synthesis.