From the School of Molecular Biosciences, Washington State University, Pullman, Washington 99164-7520.
J Biol Chem. 2012 Aug 24;287(35):29543-53. doi: 10.1074/jbc.M112.352302. Epub 2012 Jul 7.
Gene conversion is one of the frequent end results of homologous recombination, and it often underlies the inactivation of tumor suppressor genes in cancer cells. Here, we have developed an integrated assay system that allows simultaneous examination of double-strand break (DSB)-induced gene conversion events at the site of a DSB (proximal region) and at a surrounding region ~1 kb away from the break (distal region). Utilizing this assay system, we find that gene conversion events at the proximal and distal regions are relatively independent of one another. The results also indicate that synthesis-dependent strand annealing (SDSA) plays a major role in DSB-induced gene conversion. In addition, our current study has demonstrated that hMLH1 plays an essential role in anti-recombination and gene conversion. Specifically, the anti-recombination activity of hMLH1 is partially dependent on its interaction with hMRE11. Our data suggests that the role of hMLH1 and hMRE11 in the process of gene conversion is complex, and these proteins play different roles in DSB-induced proximal and distal gene conversions. In particular, the involvement of hMLH1 and hMRE11 in the distal gene conversion requires both hMSH2 and heteroduplex formation.
基因转换是同源重组的常见结果之一，它常常导致癌细胞中肿瘤抑制基因的失活。在这里，我们开发了一种集成的检测系统，可以同时检测双链断裂（DSB）诱导的基因转换事件在 DSB 位点（近端区域）和离断裂约 1kb 的周围区域（远端区域）。利用该检测系统，我们发现近端和远端区域的基因转换事件彼此相对独立。结果还表明，合成依赖性链退火（SDSA）在 DSB 诱导的基因转换中起主要作用。此外，我们目前的研究表明 hMLH1 在抗重组和基因转换中起着重要作用。具体而言，hMLH1 的抗重组活性部分依赖于其与 hMRE11 的相互作用。我们的数据表明，hMLH1 和 hMRE11 在基因转换过程中的作用是复杂的，这些蛋白在 DSB 诱导的近端和远端基因转换中发挥不同的作用。特别是，hMLH1 和 hMRE11 参与远端基因转换需要 hMSH2 和异源双链体的形成。
