Nucleotide excision repair (NER) is the primary pathway for the removal of DNA adducts that distort the double helix. In the yeast Saccharomyces cerevisiae the RAD6 epistasis group defines a more poorly characterized set of DNA damage response pathways, believed to be distinct from NER. Here we show that the elimination of the DNA minor groove adducts formed by an important class of anticancer antibiotic (CC-1065 family) requires NER factors in S. cerevisiae. We also demonstrate that the elimination of this class of minor groove adduct from the active MFA2 gene depends upon functional Rad18 and Rad6. This is most clear for the repair of adducts on the transcribed strand, where an absolute requirement for Rad6 and Rad18 was seen. Further experiments revealed that a specific RAD6-RAD18-controlled subpathway, the RAD5 branch, mediates these events. Cells disrupted for rad5 are highly sensitive to this minor groove binding agent, and rad5 cells exhibit an in vivo adduct elimination defect indistinguishable from that seen in rad6 and rad18 cells as well as in NER-defective cells. Our results indicate that the RAD5 subpathway may interact with NER factors during the repair of certain DNA adducts.