Untersuchungen zur Retrotransposition der mobilen Elemente TRE5-A und DGLT-A Dictyostelium discoideum
The haploid genome of the social amoeba Dictyostelium discoideum is characterized by a high gene density. A significant part of this genome consists of mobile genetic elements. These elements pose a high risk for insertional mutations. It is supposed that the integration of the retrotransposons TRE5-A and DGLT-A upstream of tRNA genes represents a successful strategy to avoid insertional mutations. Therefore the molecular mechanism of targeting tRNA genes through TRE5-A and DGLT-A was examined. Recent in vitro studies indicate that TRE5-A detects its integration site on the basis of a protein interaction between its encoded ORF1 protein and the transcription factor IIIB subunit TBP. Generation of TBP knock-in mutants showed that the tRNA gene directed retrotransposition of TRE5-A is based on an interaction between the ORF1 protein and helix H2’ of TBP in vivo, too. Furthermore it could be shown that a secondary structure of the 3’ end of TRE5-A RNA is essential for retrotransposition in vivo. The formation of RNA as a requirement for the retrotransposition process of TRE5-A is controlled by the host factor CbfA. By generating an in vivo retrotransposition model for TRE5-A it could be shown that CbfA has an effect on both accumulation of TRE5-A transcripts and TRE5-A retrotransposition. For the first time it was investigated whether DGLT-A targets tRNA genes by using protein interactions with subunits of transcription factors as well. Regarding this, a direct interaction between the ribonuclease H domain of DGLT-A and the subunit Tfc4 of the transcription factor IIIC could be revealed. In summary, it can therefore be said that the targeting of tRNA genes as save integration sites is independently controlled in different ways by TRE5-A and DGLT-A in D. discoideum. Consequently these data support the hypothesis that a form of convergent evolution at molecular level could help to keep mobile elements in compact genomes through integration near tRNA genes.