Brown Maxwell W, Kim Yoori, Williams Gregory M, Huck John D, Surtees Jennifer A, Finkelstein Ilya J
Department of Molecular Biosciences, Institute for Cellular and Molecular Biology, The University of Texas at Austin, Austin, Texas 78712, USA.
Department of Biochemistry, School of Medicine and Biomedical Sciences, State University of New York at Buffalo, Buffalo, New York 14214, USA.
Nat Commun. 2016 Feb 3;7:10607. doi: 10.1038/ncomms10607.
DNA-binding proteins search for specific targets via facilitated diffusion along a crowded genome. However, little is known about how crowded DNA modulates facilitated diffusion and target recognition. Here we use DNA curtains and single-molecule fluorescence imaging to investigate how Msh2-Msh3, a eukaryotic mismatch repair complex, navigates on crowded DNA. Msh2-Msh3 hops over nucleosomes and other protein roadblocks, but maintains sufficient contact with DNA to recognize a single lesion. In contrast, Msh2-Msh6 slides without hopping and is largely blocked by protein roadblocks. Remarkably, the Msh3-specific mispair-binding domain (MBD) licences a chimeric Msh2-Msh6(3MBD) to bypass nucleosomes. Our studies contrast how Msh2-Msh3 and Msh2-Msh6 navigate on a crowded genome and suggest how Msh2-Msh3 locates DNA lesions outside of replication-coupled repair. These results also provide insights into how DNA repair factors search for DNA lesions in the context of chromatin.
DNA结合蛋白通过沿着拥挤的基因组进行易化扩散来寻找特定靶点。然而，关于拥挤的DNA如何调节易化扩散和靶点识别，我们知之甚少。在这里，我们使用DNA帘和单分子荧光成像技术来研究真核生物错配修复复合物Msh2-Msh3如何在拥挤的DNA上导航。Msh2-Msh3会跳过核小体和其他蛋白质障碍，但与DNA保持足够的接触以识别单个损伤。相比之下，Msh2-Msh6则滑动而不跳跃，并且在很大程度上被蛋白质障碍所阻挡。值得注意的是，Msh3特异性错配结合结构域（MBD）使嵌合的Msh2-Msh6(3MBD)能够绕过核小体。我们的研究对比了Msh2-Msh3和Msh2-Msh6在拥挤基因组上的导航方式，并揭示了Msh2-Msh3如何在复制偶联修复之外定位DNA损伤。这些结果也为DNA修复因子如何在染色质环境中寻找DNA损伤提供了见解。
