DNA polymerase beta can incorporate ribonucleotides during DNA synthesis of undamaged and CPD-damaged DNA.
Overexpression of the error-prone DNA polymerase beta (Pol beta) has been found to increase spontaneous mutagenesis by competing with the replicative polymerases during DNA replication. Here, we investigate an additional mechanism potentially used by Pol beta to enhance genetic instability via its ability to incorporate ribonucleotides into DNA. By using an in vitro primer extension assay, we show that purified human and calf thymus Pol beta can synthesize up to 8-mer long RNA. Moreover, Pol beta can efficiently incorporate rCTP opposite G in the absence of dCTP and, to a lesser extent, rATP opposite T in the absence of dATP and rGTP opposite C in the absence of dGTP. Recently, Pol beta was shown to catalyze in vitro translesion replication of a thymine cyclobutane pyrimidine dimer (CPD). Here, we investigate if ribonucleotides could be incorporated opposite the CPD damage and modulate the efficiency of the bypass process. We find that all four rNTPs can be incorporated opposite the CPD lesion, and that this process affects translesion synthesis. We discuss how incorporation of ribonucleotides into DNA may contribute to the high frequency of mutagenesis observed in Pol beta up-regulating cells.
Bergoglio
Valérie
V
Group "Genetic Instability and Cancer" "Equipe labellisée par La Ligue Nationale contre le Cancer", Institut de Pharmacologie et Biologie Structurale, UMR CNRS 5089, 205 route de Narbonne, 31077 4, Toulouse cédex, France.
Ferrari
Elena
E
Hübscher
Ulrich
U
Cazaux
Christophe
C
Hoffmann
Jean Sébastien
JS
D000818:Animals; D001483:Base Sequence; D002417:Cattle; D004247:DNA / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D018736:DNA Adducts / Q000096:biosynthesis; D004249:DNA Damage; D019951:DNA Polymerase beta / Q000378:metabolism*; D004261:DNA Replication; D006801:Humans; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D016296:Mutagenesis; D011740:Pyrimidine Dimers / Q000378:metabolism; D012265:Ribonucleotides / Q000378:metabolism*
D018736:DNA Adducts; D011740:Pyrimidine Dimers; D012265:Ribonucleotides; D004247:DNA; D019951:DNA Polymerase beta