Department of Biological Chemistry, Johns Hopkins Medical School, Baltimore, Maryland 21205, USA.
J Biol Chem. 2010 Mar 5;285(10):7056-66. doi: 10.1074/jbc.M109.084038. Epub 2009 Dec 30.
Kinetoplast DNA, the trypanosome mitochondrial genome, is a network of interlocked DNA rings including several thousand minicircles and a few dozen maxicircles. Minicircles replicate after release from the network, and their progeny reattach. Remarkably, trypanosomes have six mitochondrial DNA helicases related to yeast PIF1 helicase. Here we report that one of the six, TbPIF1, functions in minicircle replication. RNA interference (RNAi) of TbPIF1 causes a growth defect and kinetoplast DNA loss. Minicircle replication intermediates decrease during RNAi, and there is an accumulation of multiply interlocked, covalently closed minicircle dimers (fraction U). In studying the significance of fraction U, we found that this species also accumulates during RNAi of mitochondrial topoisomerase II. These data indicate that one function of TbPIF1 is an involvement, together with topoisomerase II, in the segregation of minicircle progeny.
动基体 DNA 是锥虫线粒体基因组，是由数千个小型环和少数几十个大型环组成的连锁 DNA 环网络。小型环从网络中释放后进行复制，其后代重新附着。值得注意的是，锥虫有六个与酵母 PIF1 解旋酶相关的线粒体 DNA 解旋酶。在这里，我们报告其中一种，TbPIF1，在小型环复制中起作用。TbPIF1 的 RNA 干扰 (RNAi) 会导致生长缺陷和动基体 DNA 丢失。RNAi 过程中小型环复制中间体减少，并且会积累大量连锁的、共价闭合的小型环二聚体（U 型分数）。在研究 U 型分数的意义时，我们发现该物质也会在线粒体拓扑异构酶 II 的 RNAi 过程中积累。这些数据表明，TbPIF1 的一个功能是与拓扑异构酶 II 一起参与小型环后代的分离。
