DNA-POLYMERASE-BETA BYPASSES IN-VITRO A SINGLE D(GPG)-CISPLATIN ADDUCT PLACED ON CODON-13 OF THE HRAS GENE

We have examined the capacity of calf thymus DNA polymerases alpha, be ta, delta, and epsilon to perform in vitro translesion synthesis on a substrate containing a single d(GpG)-cisplatin adduct placed on codon 13 of the human HRAS gene, We found that DNA synthesis catalyzed by DN A. polymerases alpha, delta, and epsilon was blocked at the base prece ding the lesion, Addition of proliferating cell nuclear antigen to DNA polymerase delta and replication protein A to DNA polymerase a did no t restore their capacity to elongate past the adduct, On the other han d, DNA polymerase beta efficiently bypassed the cisplatin adduct, Furt hermore, we observed that DNA polymerase beta was the only polymerase capable of primer extension of a 3'-OH located opposite the base prece ding the lesion, Likewise, DNA polymerase beta was able to elongate th e arrested replication products of the other three DNA polymerases, th us showing its capacity to successfully compete with polymerases alpha , delta, and epsilon in the stalled replication complex, Our data sugg est (i) a possible mechanism enabling DNA polymerase beta to by-pass a d(GpG)-cisplatin adduct in vitro and (ii) a role for this enzyme in p rocessing DNA damage in vivo.