Prasad Tekkatte Krishnamurthy, Robertson Ragan B, Visnapuu Mari-Liis, Chi Peter, Sung Patrick, Greene Eric C
Department of Biochemistry and Molecular Biophysics, Columbia University, 650 West 168th Street, New York, NY 10032, USA.
J Mol Biol. 2007 Jun 15;369(4):940-53. doi: 10.1016/j.jmb.2007.04.005. Epub 2007 Apr 5.
We have used total internal reflection fluorescence microscopy (TIRFM) to investigate the characteristics of the yeast homologous recombination factor Rdh54 on DNA. Our results demonstrate translocation of Rdh54 on DNA and extrusion of DNA loops by Rdh54 in an ATP hydrolysis-dependent manner. The translocating Rdh54 was highly processive and displayed a variety of behavior, including variations in translocation rate and distance, pauses, and reversals. We provide evidence that the DNA loops generated encompass an average of 6 kb, and Rdh54 often abruptly releases the extruded DNA. Rdh54 forms a multimeric complex, which we speculate has at least two functionally distinct DNA-binding sites, one of which enables translocation while the other remains anchored to another DNA locale. Our work, together with other recent studies, suggests that translocation-coupled DNA loop extrusion is a common mechanistic feature among the Snf2-family of chromatin-remodeling proteins.
我们利用全内反射荧光显微镜（TIRFM）研究了酵母同源重组因子Rdh54在DNA上的特性。我们的结果表明，Rdh54以ATP水解依赖的方式在DNA上发生易位，并挤出DNA环。易位的Rdh54具有高度的持续性，并表现出多种行为，包括易位速率和距离的变化、停顿和反转。我们提供的证据表明，产生的DNA环平均包含6 kb，并且Rdh54经常突然释放挤出的DNA。Rdh54形成多聚体复合物，我们推测其至少有两个功能不同的DNA结合位点，其中一个使易位发生，而另一个则锚定在另一个DNA区域。我们的工作与最近的其他研究一起表明，易位偶联的DNA环挤出是染色质重塑蛋白Snf2家族中的一个常见机制特征。
