Protein Interactions in Antibody Diversity

DNA聚合酶IOTA(Poliota)因其保真度低而成为抗体基因体细胞高突变的候选基因。为了确定Pol iota的作用，我们分析了两种类型的缺乏酶的小鼠的突变：129只表达截短的Pol iota的小鼠，以及表达全长Pol iota的敲击小鼠，该全长Pol iota是催化不活跃的。两个菌株在可变区的突变频率和光谱都是正常的，这表明Pol iota的丢失并没有改变整体的突变。然后，我们检查了Pol iota是否影响了另一种容易出错的聚合酶Pol Zeta产生的串联突变。使用一种新的计算模型分析了连续突变的频率，以确定它们是发生在单个DNA交易期间还是发生在两个独立事件期间。对两个菌株的2000个突变的分析表明，Pol iota受损的小鼠失去了串联签名，而C57BL/6小鼠积累了大量的双重突变。结果支持这样一种模型，即Pol iota偶尔访问复制分叉以产生第一个突变，而Pol Zeta通过第二个突变延长不匹配。

DNA polymerase iota (Pol iota) is an attractive candidate for somatic hypermutation in antibody genes because of its low fidelity. To identify a role for Pol iota, we analyzed mutations in two strains of mice with deficiencies in the enzyme: 129 mice with negligible expression of truncated Pol iota, and knockin mice which express full length Pol iota that is catalytically inactive. Both strains had normal frequencies and spectra of mutations in the variable region, indicating that loss of Pol iota did not change overall mutagenesis. We then examined if Pol iota affected tandem mutations generated by another error prone polymerase, Pol zeta. The frequency of contiguous mutations was analyzed using a novel computational model to determine if they occur during a single DNA transaction or during two independent events. Analyses of 2,000 mutations from both strains indicated that Pol iota-compromised mice lost the tandem signature, whereas C57BL/6 mice accumulated significant amounts of double mutations. The results support a model where Pol iota occasionally accesses the replication fork to generate a first mutation, and Pol zeta extends the mismatch with a second mutation.