Transcription-coupled nucleotide excision repair (TCR) accelerates the removal 
of noncoding lesions from the template strand of active genes, and hence 
contributes to genome-wide variations in mutation frequency. Current models for 
TCR suppose that a lesion must cause RNA polymerase (RNAP) to stall if it is to 
be a substrate for accelerated repair. We have examined the substrate 
requirements for TCR using a system in which transcription stalling and damage 
location can be uncoupled. We show that Mfd-dependent TCR in bacteria involves 
the formation of a damage search complex that can detect lesions downstream of a 
stalled RNAP, and that the strand specificity of the accelerated repair pathway 
is independent of the requirement for a lesion to stall RNAP. We also show that 
an ops (operon polarity suppressor) transcription pause site, which causes 
backtracking of RNAP, can promote the repair of downstream lesions when those 
lesions do not themselves cause the polymerase to stall. Our findings indicate 
that the transcription-repair coupling factor Mfd, which is an ATP-dependent 
superfamily 2 helicase that binds to RNAP, continues to translocate along DNA 
after RNAP has been displaced until a lesion in the template strand is located. 
The discovery that pause sites can promote the repair of nonstalling lesions 
suggests that TCR pathways may play a wider role in modulating mutation 
frequencies in different parts of the genome than has previously been suspected.