FOR 1082: Biochemistry and Biological Function of Dnmt2 Methyltransferases

点击翻译按钮获取中文摘要

Proteins of the DNA methyltransferase (Dnmt) family play a pivotal role in epigenetic gene regulation. Research in this area has and will have a strong impact on our understanding of gene regulation, organismal development and human disease. Within the Dnmt family, the roles of Dnmt1 (maintenance methyltransferase) and Dnmt3 (de novo methyltransferases) are relatively well understood. In contrast, Dnmt2 proteins have remained rather enigmatic. This is mostly due to a severely limited understanding of their biochemistry and the poor definition of their biological function. Among all Dnmt proteins, Dnmt2 is the most highly conserved and homologues have been described in organisms as diverse as protists (Entamoeba, Dictyostelium), plants, protostomia and mammals. Phenotypes of Dnmt2 knock-outs or knock-downs vary from severe (zebrafish) to barely detectable (Drosophila, Dictyostelium, Schizosaccharomyces pombe, mouse). Although the enzyme shares all amino acid motifs characteristic for DNA methyltransferases, only a weak DNA-methylation activity of Dnmt2 could be shown in vitro. However, there is strong experimental evidence that some genomic loci, in particular retroelements, are metylated in vivo by Dnmt2. More recently, a second activity, the site-specific methylation of a tRNA has been shown. The Research Unit aims at a comparative biochemical analysis of Dnmt2 proteins from human, Drosophila, Entamoeba, Dictyostelium and S. pombe to obtain insights into the enzymatic function on DNA as well as on RNA. This includes the definition of substrates as well as properties of the enzyme that are required to recognise and modify the two different substrates. It is likely that cofactors are required for enzyme activity, the Research Unit will, therefore, identitify proteins that interact with Dnmt2 and isolate synthetic lethal mutants in yeast. To specify further targets, co-immune precipitations will be done and associated DNA and RNA molecules will be identified. The effect of Dnmt2 on tRNA activity in translation systems in vivo and in vitro is investigated as well as the significance in silencing and mobility of retroelements. In vivo readout systems have been designed in order to understand the biological function of Dnmt2.