Initiation of DNA repair mediated by a stalled RNA polymerase IIO

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Abstract
The transcription‐coupled repair (TCR) pathway preferentially repairs DNA damage located in the transcribed strand of an active gene. To gain insight into the coupling mechanism between transcription and repair, we have set up an in vitro system in which we isolate an elongating RNA pol IIO, which is stalled in front of a cisplatin adduct. This immobilized RNA pol IIO is used as ‘bait’ to sequentially recruit TFIIH, XPA, RPA, XPG and XPF repair factors in an ATP‐dependent manner. This RNA pol IIO/repair complex allows the ATP‐dependent removal of the lesion only in the presence of CSB, while the latter does not promote dual incision in an XPC‐dependent nucleotide excision repair reaction. In parallel to the dual incision, the repair factors also allow the partial release of RNA pol IIO. In this ‘minimal TCR system’, the RNA pol IIO can effectively act as a loading point for all the repair factors required to eliminate a transcription‐blocking lesion.