Altered nucleotide misinsertion fidelity associated with poliota-dependent replication at the end of a DNA template

Abstract

A hallmark of human DNA polymerase ι (polι) is the asymmetric fidelity of replication at template A and T when the enzyme extends primers annealed to a single‐stranded template. Here, we report on the efficiency and accuracy of polι‐dependent replication at a nick, a gap, the very end of a template and from a mispaired primer. Polι cannot initiate synthesis on a nicked DNA substrate, but fills short gaps efficiently. Surprisingly, polι9s ability to blunt‐end a 1 bp recessed terminus is dependent upon the template nucleotide encountered and is highly erroneous. At template G, both C and T are inserted with roughly equal efficiency, whilst at template C, C and A are misinserted 8‐ and 3‐fold more often than the correct base, G. Using substrates containing mispaired primer termini, we show that polι can extend all 12 mispairs, but with differing efficiencies. Polι can also extend a tandem mispair, especially when it is located within a short gap. The enzymatic properties of polι appear consistent with that of a somatic hypermutase and suggest that polι may be one of the low‐fidelity DNA polymerases hypothesized to participate in the hypermutation of immunoglobulin variable genes in vivo.