Department of Microbiology, University of Georgia, Athens, Georgia, USA.
J Bacteriol. 2012 Aug;194(16):4395-405. doi: 10.1128/JB.00783-12. Epub 2012 Jun 15.
Recombination between insertion sequence copies can cause genetic deletion, inversion, or duplication. However, it is difficult to assess the fraction of all genomic rearrangements that involve insertion sequences. In previous gene duplication and amplification studies of Acinetobacter baylyi ADP1, an insertion sequence was evident in approximately 2% of the characterized duplication sites. Gene amplification occurs frequently in all organisms and has a significant impact on evolution, adaptation, drug resistance, cancer, and various disorders. To understand the molecular details of this important process, a previously developed system was used to analyze gene amplification in selected mutants. The current study focused on amplification events in two chromosomal regions that are near one of six copies of the only transposable element in ADP1, IS1236 (an IS3 family member). Twenty-one independent mutants were analyzed, and in contrast to previous studies of a different chromosomal region, IS1236 was involved in 86% of these events. IS1236-mediated amplification could occur through homologous recombination between insertion sequences on both sides of a duplicated region. However, this mechanism presupposes that transposition generates an appropriately positioned additional copy of IS1236. To evaluate this possibility, PCR and Southern hybridization were used to determine the chromosomal configurations of amplification mutants involving IS1236. Surprisingly, the genomic patterns were inconsistent with the hypothesis that intramolecular homologous recombination occurred between insertion sequences following an initial transposition event. These results raise a novel possibility that the gene amplification events near the IS1236 elements arise from illegitimate recombination involving transposase-mediated DNA cleavage.
插入序列之间的重组可能导致遗传缺失、倒位或重复。然而，要评估涉及插入序列的所有基因组重排的比例是很困难的。在先前对不动杆菌 ADP1 的基因重复和扩增研究中，在大约 2%的特征重复位点中发现了一个插入序列。基因扩增在所有生物体中都很常见，对进化、适应、耐药性、癌症和各种疾病都有重大影响。为了了解这个重要过程的分子细节，先前开发的系统被用于分析选定突变体中的基因扩增。本研究集中在两个染色体区域的扩增事件上，这两个区域靠近 ADP1 中唯一的转座元件 IS1236（IS3 家族成员）的六个拷贝之一。对 21 个独立的突变体进行了分析，与之前对不同染色体区域的研究相比，IS1236 参与了其中 86%的事件。IS1236 介导的扩增可以通过重复区域两侧插入序列之间的同源重组发生。然而，这种机制假定转座产生了一个位置适当的额外 IS1236 拷贝。为了评估这种可能性，使用 PCR 和 Southern 杂交来确定涉及 IS1236 的扩增突变体的染色体构型。令人惊讶的是，基因组模式与以下假设不一致：在初始转座事件之后，插入序列之间的分子内同源重组发生。这些结果提出了一个新的可能性，即在 IS1236 元件附近的基因扩增事件源自涉及转座酶介导的 DNA 切割的非法重组。
