Mammalian L1 retrotransposons as genetic characters

最新发现 使用L1插入来确定影响突变率的因素-中性DNA突变率(即在没有自然选择的情况下碱基替换的积累)是一个基本的生物学参数。有趣的是，它在染色体内和染色体之间有所不同，但尽管进行了大量研究，这种差异在很大程度上仍然无法解释。例如，中性突变率随着CpG含量的不同而变化，这种相关性被认为反映了两者都是基因组环境的协变量的事实。然而，通过比较不同L1化石的突变率，我们发现CPGS本身或其突变可以直接影响侧翼非CpG DNA的突变(Walser等人，Genome 18：1403；Walser&Furano，Genome 20：875)。这项工作的一个重要结果是，足以影响非CpG分歧的CpG含量也会影响突变环境，产生的转换/颠换比率让人想起一些肿瘤的“突变子表型”。因此，我们实施了一个实验系统，直接在体内确定DNA修复是否可以诱导侧翼DNA发生突变，并发现确实如此。特别是，在基于SV40的Episome上，由预先形成的正常发生的DNA错配产生的修复中间产物在低频率但在统计上显著地容易受到APOBEC介导的易于出错的过程的影响。SiRNA敲除表明，碱基切除修复和错配修复途径的组件，或可以与这些途径相互作用的因子(例如，增殖细胞核抗原和ATR)，以及TPC偏好的APOBEC脱氨酶都是突变所必需的，这种突变产生的突变类似于各种癌症中典型的突变子表型。因此，正常情况下，没有错误的DNA修复过程可以转变为突变，提供了迄今为止意想不到的基因变化来源，这些变化是疾病、衰老和进化变化的基础。我们现在已经提交了这些结果以供发布。

CURRENT FINDINGS USING L1 INSERTS TO DETERMINE THE FACTORS THAT AFFECT MUTATION RATE - The neutral DNA mutation rate (i.e., the accumulation of base substitutions in the absence of natural selection) is a fundamental biological parameter. Interestingly, it varies within and between chromosomes, but despite considerable study, this variation remains largely unexplained. For example, the neutral mutation rate varies with CpG content, a correlation thought to reflect the fact that the both are covariates of the genomic environment. However, by comparing the mutation rates of different L1 fossils we showed that CpGs per se, or mutations thereof, can directly affect the mutation of flanking non-CpG DNA( Walser, et al, Genome Res 18: 1403; Walser & Furano, Genome Res 20: 875). And an important result of this work was that a CpG content sufficient to affect non-CpG divergence also affects the mutational environment, producing transition/transversion ratios that are reminiscent of the "mutator phenotype" of some tumors. We therefore implemented an experimental system to directly determine in vivo whether DNA repair can induce mutations in flanking DNA and found that it does. In particular, the repair intermediates generated from preformed normally occurring DNA mispairs on an SV40-based episome were vulnerable at a low but statistically significant frequency to an APOBEC-mediated error-prone process. SiRNA knockdowns showed that components of both the base excision repair and mismatch repair pathways, or factors that can interact with these pathways (e.g., PCNA and ATR), and TpC-preferring APOBEC deaminases are all required for mutagenesis which produces mutations similar to those typical of the mutator phenotypes in various cancers. Thus normally error-free DNA repair processes can be turned into mutators providing a heretofore unexpected source of genetic changes that underlie disease, aging and evolutionary change. We have now submitted these results for publication.