Structures and mechanism of transcription initiation by bacterial ECF factors

Bacterial RNA polymerase (RNAP) forms distinct holoenzymes with extra-cytoplasmic function (ECF) σ factors to initiate specific gene expression programs. In this study, we report a cryo-EM structure at 4.0 Å of Escherichia coli transcription initiation complex comprising σ(E)—the most-studied bacterial ECF σ factor (Ec σ(E)-RPo), and a crystal structure at 3.1 Å of Mycobacterium tuberculosis transcription initiation complex with a chimeric σ(H/E) (Mtb σ(H/E)-RPo). The structure of Ec σ(E)-RPo reveals key interactions essential for assembly of E. coli σ(E)-RNAP holoenzyme and for promoter recognition and unwinding by E. coli σ(E). Moreover, both structures show that the non-conserved linkers (σ(2)/σ(4) linker) of the two ECF σ factors are inserted into the active-center cleft and exit through the RNA-exit channel. We performed secondary-structure prediction of 27,670 ECF σ factors and find that their non-conserved linkers probably reach into and exit from RNAP active-center cleft in a similar manner. Further biochemical results suggest that such σ(2)/σ(4) linker plays an important role in RPo formation, abortive production and promoter escape during ECF σ factors-mediated transcription initiation.