DNA protein crosslinks (DPCs) can form when cellular proteins are covalently trapped on DNA following exposure to chemotherapeutic agents, UV radiation, reactive oxygen species (ROS), transition metals, and aldehydes. The bulky lesions that form as a result can inhibit DNA repair, replication, and transcription. 5-Formyl cytosine (5fC) is an epigenetic modification to DNA containing a reactive aldehyde group that reacts with H2A and H4 histone proteins, amongst others. These DPCs can undergo proteolytic cleavage to produce DNA-peptide conjugates. The Tretyakova lab has shown that while DNA-protein crosslinks block replication, DNA-peptide crosslinks can be bypassed by human translesion synthesis (TLS) polymerases η and κ in an error-prone manner both <i>in vitro </i>and in human cells. However, the role of the DNA sequence context in DNA replication of the DNA-peptide crosslinks has not been studied. In the present study, 5fC-containing DNA of varying sequences were crosslinked to an 11-mer peptide via an oxime ligation method using oxo-lysine. The bypass of DNA-peptide crosslinks by TLS human polymerase η <i>in vitro</i> was investigated, including the steady state kinetics of incorporation of correct and incorrect base addition opposite the DPC lesion. We observed significant differences in the efficiency of nucleotide incorporation when the 5’ and/or 3’ base to the lesion site is altered. In an additional study, lesion bypass by replicative human polymerases δ and ε was investigated, which showed that these replicative polymerases can bypass the DNA peptide lesions in an error-free manner. Taken together, these results indicate that the effects of DNA-peptide cross-links of DNA replication are strongly dependent on DNA sequence context and the identity of DNA polymerase.

Translesion synthesis past DNA-peptide crosslinks is dependent on local DNA sequence context

2019 Fall National Meeting

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