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TitleSingle-stranded DNA binding to the transcription factor PafBC triggers the mycobacterial DNA damage response.
Journal, issue, pagesSci Adv, Vol. 11, Issue 6, Page eadq9054, Year 2025
Publish dateFeb 7, 2025
AuthorsCharlotte M Schilling / Rafal Zdanowicz / Julius Rabl / Andreas U Müller / Daniel Boehringer / Rudi Glockshuber / Eilika Weber-Ban /
PubMed AbstractThe DNA damage response in mycobacteria is controlled by the heterodimeric transcription factor PafBC, a member of the WYL domain-containing protein family. It has been shown that PafBC induces ...The DNA damage response in mycobacteria is controlled by the heterodimeric transcription factor PafBC, a member of the WYL domain-containing protein family. It has been shown that PafBC induces transcription of its regulon by reprogramming the housekeeping RNA polymerase holoenzyme to recognize PafBC-dependent promoters through sigma adaptation. However, the mechanism by which DNA damage is sensed and translated into PafBC activation has remained unclear. Here, we demonstrate that the binding of single-stranded DNA (ssDNA) to the WYL domains of PafBC activates the transcription factor. Our cryo-electron microscopy structure of full-length PafBC in its active conformation, bound to the transcription initiation complex, reveals a previously unknown mode of interaction between the ssDNA and the WYL domains. Using biochemical experiments, we show that short ssDNA fragments bind to PafBC dynamically, resulting in deactivation as ssDNA levels decrease postrepair. Our findings shed light on the mechanism linking DNA damage to PafBC activation and expand our understanding of WYL domain-containing proteins.
External linksSci Adv / PubMed:39919186 / PubMed Central
MethodsEM (single particle)
Resolution3.3 - 4.0 Å
Structure data

EMDB-50589: DNA-directed RNA polymerase with transcriptional activator PafBC
Method: EM (single particle) / Resolution: 3.3 Å

50590

9fnd

EMDB-50590, PDB-9fnd:
Transcriptional activator PafBC bound to mycobacterial RNA polymerase
Method: EM (single particle) / Resolution: 4.0 Å

50591

9fne

EMDB-50591: Composite map of the mycobacterial PafBC-bound transcription initiation complex
PDB-9fne: Mycobacterial PafBC-bound transcription initiation complex
Method: EM (single particle) / Resolution: 4.0 Å

Chemicals

ChemComp-ZN:
Unknown entry

ChemComp-MG:
Unknown entry

Source
  • mycolicibacterium smegmatis mc2 155 (bacteria)
KeywordsTRANSCRIPTION / PafBC / sigma adaptation / WYL domain / transcriptional activator / RNAP

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