Functional characterization of Dnmt2 in Drosophila

dnmt 2酶代表了理解胞嘧啶-5甲基化的生物学功能的中心范式，胞嘧啶-5甲基化是一种与许多人类疾病（包括癌症）相关的核酸修饰。Dnmt 2利用DNA甲基转移酶的催化机制，但仅在tRNA底物上观察到稳健的催化活性。这使得Dnmt 2酶非常适合研究RNA和DNA中胞嘧啶甲基化的生物学功能。我们将专注于Dnmt 2在果蝇和小鼠中的表征，因为果蝇已被证明是非常适合结合分子，生物化学和遗传学的方法，而小鼠模型将这些观察与哺乳动物生物学。我们以前的研究结果使我们能够建立Dnmt 2作为一种多底物tRNA甲基转移酶，并确定这些酶在细胞应激反应中的作用。我们现在将（1）分析Dnmt 2依赖性tRNA片段在应激信号传导中的功能，（2）确定Dnmt 2与小RNA沉默途径的相互作用，（3）表征胞嘧啶-5 tRNA甲基化缺陷小鼠模型的分化表型，以及（4）在全转录组水平上确定胞嘧啶-5 RNA甲基化模式。

Dnmt2 enzymes represent a central paradigm for understanding the biological function of cytosine-5 methylation, a nucleic acid modification that has been associated with numerous human diseases, including cancer. Dnmt2 utilizes the catalytic mechanism of DNA methyltransferases, but robust catalytic activity has only been observed at tRNA substrates. This makes Dnmt2 enzymes well suited to investigate the biological function of cytosine methylation in RNA and DNA. We will focus on the characterization of Dnmt2 in Drosophila and in the mouse, because Drosophila has been proven to be highly suitable for combined molecular, biochemical and genetic approaches whereas the mouse model links these observations to mammalian biology. Our previous findings allowed us to establish Dnmt2 as a multi-substrate tRNA methyltransferase and to identify a role of these enzymes in cellular stress responses. We will now (1) analyze the function of Dnmt2-dependent tRNA fragmentation in stress signaling, (2) determine the interaction of Dnmt2 with small RNA silencing pathways, (3) characterize differentiation phenotypes of cytosine-5 tRNA methylation-deficient mouse models and (4) determine cytosine-5 RNA methylation patterns on the whole-transcriptome level.