Characterization of E.histolytica DNMT2 homolog EhDNMT2 and the meaning of its interaction with Enolase

在资助的第一个阶段，我们已经证明了短期葡萄糖饥饿通过烯醇化酶（一种糖酵解酶）与Ehmeth的结合来调节Ehmeth的活性。我们将集中研究肌钙蛋白的表征，这是最近发现的另一种Dnmt2相互作用蛋白。与烯醇化酶的行为相反，在葡萄糖饥饿的寄生虫中，肌钙蛋白与Ehmeth的相互作用较少。这种相互作用的意义仍然是未知的，它将通过使用生物化学和分子生物学技术的组合在这个联盟的框架内进行研究。接下来，我们将探讨长期葡萄糖饥饿、一氧化氮和热休克等其他环境因素对Ehmeth表达及其活性的影响。我们还将更详细地研究逆转录因子在激活Ehmeth表达中的作用。为了理解为什么Ehmeth在我们的实验室菌株中表达如此之低，我们将描述其启动子区域和调控元件的存在。这种全球方法将帮助我们了解营养和环境胁迫对Dnmt2表达的作用，这一现象也在该联盟研究的生物体中观察到。

During the first period of funding, we have demonstrated that short term glucose starvation regulates Ehmeth activity via the binding of enolase, a glycolytic enzyme to Ehmeth. We will concentrate our research on the characterization of troponin, a second Dnmt2 interacting protein that has been recently identified. In contrast to the behavior of enolase, less troponin interacts with Ehmeth in glucose starved parasites. The meaning of this interaction is still unknown and it will be studied within the framework of this consortium by using a combination of biochemical and molecular biology techniques. Next, we would like to address the influence of other environmental factors including long term glucose starvation, nitric oxide and heat shock, on the expression of Ehmeth and on its activity. We will also investigate more in details the role of retroelements in the activation of Ehmeth expression. In an effort to understand why Ehmeth expression is so low in our laboratory strain, we will characterize its promoter region and the presence of regulatory elements. This global approach will help us to understand the role of nutritional and environmental stresses on Dnmt2 expression, a phenomenon that has also been observed among the organisms studied in this consortium.