Abstact

DNA damage repair mechanisms are vital for cell survival. In the bacterium, Caulobacter crescentus, DriD is the master regulator of a unique, noncanonical DNA damage pathway. DriD binding to ssDNA, produced upon DNA damage, stimulates its ability to activate transcription from several promoters involved in DNA damage responses. However, the mechanism by which DriD interfaces with the RNAP holoenzyme to activate transcription from its multiple promoters has been unclear. Here, we describe cryo-EM structures of DriD-ssDNA bound to RNAP-holoenzyme and three DriD-regulated promoters. Each subunit of homodimeric DriD contains an DNA binding N-terminal winged helix-turn-helix (wHTH) connected to WYL domains by a linker 3-helix bundle (3HB) module. The structures reveal a mechanism of assembly on promoters whereby DriD's 3HBs bind the RNAP alpha-CTD and beta domains, anchoring the RNAP-holoenzyme to regulated promoters. The 3HBs form autoinhibitory contacts with DNABDs in apo DriD and therefore acts as an ssDNA-driven trigger domain, switching between DNABD-bound apo and RNAP-bound forms upon ssDNA-mediated activation. Thus, the structures reveal a unique transcription activation mechanism, likely conserved among the large family of homodimeric WYL activators.