Human DNA polymerase has a strong mutagenic potential at the initial steps of DNA synthesis

DNA polymerase alpha (Pol alpha) is essential for DNA replication initiation and makes a notable contribution to genome mutagenesis. The activity and fidelity of Pol alpha during the early steps of DNA replication have not been well studied. Here we show that at the beginning of DNA synthesis, when extending the RNA primer received from primase, Pol alpha is more mutagenic than during the later DNA elongation steps. Kinetic and binding studies revealed substantially higher activity and affinity to the template:primer when Pol alpha interacts with ribonucleotides of a chimeric RNA-DNA primer. Pol alpha activity greatly varies during first six steps of DNA synthesis, and the bias in the rates of correct and incorrect dNTP incorporation leads to impaired fidelity, especially upon the second step of RNA primer extension. Furthermore, increased activity and stability of Pol alpha/template:primer complexes containing RNA-DNA primers result in higher efficiency of mismatch extension.