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

A hallmark of human DNA polymerase iota (pol iota) is the asymmetric fideli ty of replication at template A and T when the enzyme extends primers annea led to a single-stranded template. Here, we report on the efficiency and ac curacy of pol iota -dependent replication at a nick, a gap, the very end of a template and from a mispaired primer, Pol iota cannot initiate synthesis on a nicked DNA substrate, but fills short gaps efficiently. Surprisingly, pol iota 's ability to blunt-end a 1 bp recessed terminus is dependent upo n the template nucleotide encountered and is highly erroneous. At template G, both C and T are inserted with roughly equal efficiency, whilst at templ ate C, C and A are misinserted 8- and 3-fold more often than the correct ba se, G, Using substrates containing mispaired primer termini, we show that p ol iota can extend all 12 mispairs, but with differing efficiencies. Pol io ta can also extend a tandem mispair, especially when it is located within a short gap. The enzymatic properties of pol iota appear consistent with tha t of a somatic hypermutase and suggest that pol iota may be one of the low- fidelity DNA polymerases hypothesized to participate in the hypermutation o f immunoglobulin variable genes in vivo.