Inefficient repair of RNA center dot DNA hybrids

RNA.DNA hybrids are commonly observed during normal biological processes. We tested the ability of three DNA-repair enzymes to remove lesions from the DNA strand of RNA.DNA heteroduplexes. Three nucleotide analogs, 5-hydroxy-2'-deoxycytidine triphosphate, 8-oxo-2'-deoxyguanosine triphosphate, and O-6-methyl-2'-deoxyguanosine triphosphate, representative of lesions generated by oxygen damage and methylating agents, were incorporated into the DNA strand synthesized using either a DNA or RNA template. The extended DNA.DNA and RNA.DNA hybrids were used as substrates for bacterial formamidopyrimidine-DNA glycosylase, Nth protein (endonuclease III) and O-6-methylguanine-DNA methyltransferase. We show that all three lesions are readily cleaved from the DNA strand of a DNA.DNA duplex but are relatively resistant to cleavage when present in the DNA strand of an RNA.DNA hybrid. Our in vitro studies suggest that damaged DNA in RNA.DNA hybrids is less likely to be repaired in vivo.