Functional characterization of the Dnmt2 homolog DnmA in Dictyostelium

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In the frame of the Forschergruppe (FOR), this project will continue to obtain insights into the biochemistry and function of the methyltransferase DnmA in Dictyostelium. We will investigate the function of newly identified substrate RNAs and continue attempts to identify protein components as well as DNA sequences that associate with DnmA. Our data suggest regulation of DnmA during the cell cycle and during development. We will design experiments that will elucidate DnmA functions at these specific time points in the life cycle of a cell. The significantly higher expression levels in some wild isolates will substantially facilitate this approach. An intriguing feature of Dnmt2 is its high conservation in evolution which is contrasted by the lack of obvious mutant phenotypes. Even more surprising is the observation that KO cells appear to have a growth advantage over wild type cells in mixed cultures. We will elucidate the factors responsible for this unexpected finding. We hope to define conditions where DnmA is required and thus provide indications for its evolutionary conserved function. Specific single chain antibodies against DnmA and hDnmt2 have been obtained and will be optimised to detect the endogenous proteins. As in the previous funding period, experiments will be carried out in close cooperation with the partners in the FOR. Wherever possible, comparative studies will be done in parallel with Dnmt2 homologues from the different model organisms.