TRANSPOSASE PROTEIN METNASE IN LEUKEMIC DECATENATION

描述（由申请人提供）：转座酶活性被认为在人类中已经灭绝，因为DNA运动在高等生物中可能是有害的，导致基因组不稳定和可能的恶性肿瘤。我们分离了一种被称为Metnase的人转座酶蛋白，它对超螺旋DNA具有优先的内切酶活性。因此，我们探索了它在去除纠缠DNA中的作用。DNA复制的结果是交织在一起的姐妹染色单体，在染色单体后期分离之前必须将其解开。当缺失时，可发生灾难性的染色单体断裂，多次断裂可导致染色体间末端连接和随后的易位。因此，细胞积极地监测交织染色单体的十烷化。在有丝分裂前，链结DNA的存在通常会引起细胞周期阻滞，称为十链结检查点。先前的研究表明，膀胱癌和肺癌并不能在十烷十二周细胞周期检查点正常停止。我们还发现急性白血病不会在十位数检查点停止。当ATR检测到染色体链链时，十链链检查点被激活，然后发出信号，在有丝分裂前细胞停止。Topo II1是物理解缠染色单体所必需的脱羧酶。除此之外，人们对这种最近被描述的现象的分子机制知之甚少。我们发现Metnase不是介导DNA片段的迁移，而是通过显著提高Topo II1的DNA十烷化率来增加染色体的稳定性。Metnase水平的增加增加了对Topo II1抑制剂ICRF-193和VP-16的耐药性。基于这些基本发现，我们假设Metnase在造血十烷化中发挥作用，并通过十烷化细胞周期检查点协助进展。本应用程序将通过解决三个问题来描述这种情况发生的机制：1)Metnase增强Topo II1十烷化活性的机制是什么？2)甲基化酶的十烷基化活性是由磷酸化信号激活的吗？3) Metnase是否介导造血恶性肿瘤（如白血病）中的十烷磺酸？破译Metnase在十烷化中起作用的机制可以深入了解白血病易位的机制，为什么白血病不能在十烷化检查点适当地停止，以及一些白血病对Topo II1抑制剂的耐药性。公共卫生相关性：我们已经分离出一种称为Metnase的新蛋白质，它可以帮助染色体解开缠结，从而防止它们在细胞分裂期间断裂。防止这种染色体断裂可以减少导致多种癌症的突变。然而，癌细胞可以破坏甲基化酶，并利用它来抵抗破坏DNA的化疗。

DESCRIPTION (provided by applicant): Transposase activity was thought to be extinct in humans because DNA movement can be deleterious in higher organisms, resulting in genomic instability and perhaps malignancy. We isolated a human transposase protein termed Metnase that had preferential endonuclease activity for supercoiled DNA. We therefore explored its role in decatenating tangled DNA. DNA replication results in intertwined sister chromatids that must be untangled, or decatenated, before chromatid separation at anaphase. When absent, catastrophic chromatid breakage can occur, and the multiple breaks can result in inter- chromosomal end joining and subsequent translocation. Therefore, cells actively monitor the decatenation of intertwined chromatids. The presence of catenated DNA usually induces a cell cycle arrest, termed the decatenation check point, before mitosis. It has been previously shown that bladder and lung cancers do not properly arrest at the decatenation cell cycle checkpoints. We also found that acute leukemias do not arrest at the decatenation checkpoints. The decatenation checkpoints are activated when ATR senses catenated chromosomes and then signals for the cell to arrest before mitosis. Topo II1 is the essential decatenating enzyme for physically untangling chromatids. Beyond that, little is known about the molecular mechanism of this recently described phenomenon. We found that Metnase, rather than mediating the mobility of DNA segments, increased chromosome stability by markedly enhancing the rate of DNA decatenation by Topo II1. Increasing Metnase levels increased resistance to the Topo II1 inhibitors ICRF-193 and VP-16. Based on these fundamental findings, we hypothesize that Metnase plays a role in hematopoietic decatenation, and assists in progression through the decatenation cell cycle checkpoint. This application will characterize the mechanism by which this takes place by addressing three questions: 1) What is the mechanism by which Metnase enhances Topo II1 decatenation activity? 2) Is the decatenation activity of Metnase activated by phosphorylation signals?,3) Does Metnase mediate decatenation in hematopoietic malignancies such as leukemia? Deciphering the mechanism by which Metnase functions in decatenation could lend insight into the mechanism of leukemogenic translocations, why leukemia fails to appropriately arrest at the decatenation check points, and resistance of some leukemias to Topo II1 inhibitors. PUBLIC HEALTH RELEVANCE: We have isolated a novel protein termed Metnase that helps chromosomes untangle, and thereby prevents their breakage during cell division. Preventing such chromosome breakage can reduce the mutations that cause many forms of cancer. The cancer cell, however, can subvert Metnase, and use it to resist the actions of DNA damaging chemotherapy.