DNA polymerase ζ generates tandem mutations in immunoglobulin variable regions

Low-fidelity DNA polymerases introduce nucleotide substitutions in immunoglobulin variable regions during somatic hypermutation. Although DNA polymerase (pol) η is the major low-fidelity polymerase, other DNA polymerases may also contribute. Existing data are contradictory as to whether pol ζ is involved. We reasoned that the presence of pol η may mask the contribution of pol ζ, and therefore we generated mice deficient for pol η and heterozygous for pol ζ. The frequency and spectra of hypermutation was unaltered between Polζ(+/−) Polη(−/−) and Polζ(+/+) Polη(−/−) clones. However, there was a decrease in tandem double-base substitutions in Polζ(+/−) Polη(−/−) cells compared with Polζ(+/+) Polη(−/−) cells, suggesting that pol ζ generates tandem mutations. Contiguous mutations are consistent with the biochemical property of pol ζ to extend a mismatch with a second mutation. The presence of this unique signature implies that pol ζ contributes to mutational synthesis in vivo. Additionally, data on tandem mutations from wild type, Polζ(+/−), Polζ(−/−), Ung(−/−), Msh2(−/−), Msh6(−/−), and Ung(−/−) Msh2(−/−) clones suggest that pol ζ may function in the MSH2–MSH6 pathway.