Transcriptional bypass of bulky DNA lesions causes new mutant RNA transcripts in human cells

Here, we characterize the mutant transcripts resulting from bypass of an 8,5'-cyclo-2'-deoxyadenosine (cyclo-dA) or cyclobutane pyrimidine dimer (CPD) by human RNA polymerase II (Pol II) in vivo. With the cyclo-dA lesion, we observed two new types of mutant transcripts. In the first type, the polymerase inserted uridine opposite the lesion and then misincorporated adenosine opposite the template deoxyadenosine downstream (50) of the lesion. The second type contained deletions of 7, 13 or 21 nucleotides (nt) after uridine incorporation opposite the lesion. The frequency of the different types of transcript from the cyclo-dA lesion in mutant human cell lines suggests that the Cockayne syndrome B protein affects the probability of deletion transcript formation. With the CPD-containing construct, we also detected rare transcripts containing 12 nt deletions. These results indicate that RNA pol II in living human cells can bypass helix-distorting DNA lesions that are substrates for nucleotide excision repair, resulting in transcriptional mutagenesis.