Untersuchungen zu positionsspezifisch integrierenden Retrotransposons in sozialen Amöben
Social amoebae have very compact genomes. Therefore, it is likely that tRNA-gene-specific integration in the genome of the social amoeba Dictyostelium discoideum is an adaption to avoid insertional mutagenesis. In this study, a survey of mobile genetic elements was performed in the recently sequenced species D. purpureum, Polysphondylium pallidum and D. fasciculatum with emphasis on tRNA gene-targeting retrotransposons. Considering phylogenetic relations between different species, the detection of tRNA gene-specific elements in all species indicates even the last common ancestor of social amoebae possessed such elements within their genomes. The generation of a codon-adapted, tagged TRE5-A element enabled the discrimination of these tagged retrotransposons from the endogenous TRE5-A population. It was found that most tRNA genes of D. discoideum are principle targets for TRE5-A integration. New TRE5-A integrations were observed to occur into the extrachromosomal rDNA palindrome that carries the ribosomal RNA genes. It was shown that, distinct A- and B-box-motives on the palindrome, which have strong similarities with the features of a tRNA-gene, are recognized by the artificial TRE5-A-retrotransposons. The authentic distances of integrated TRE5-A copies to the targeted B box motifs and the 5S gene indicates an involvement of RNA-polymerase-III-transcription units in integration site recognition. It was shown that the targeted B box loci are actively transcribed, which argues for an involvement of the RNA polymerase III complex in target site selection by TRE5-A. In this work genetically tagged LTR retrotransposons were developed and characterized. It could be shown that a selection marker for retrotransposition events can be introduced into the elements without disruption of transposon activity. Interestingly, it seemed that such tagged retrotransposons were only mobilized in D. discoideum mutants with a defective RNA interference machinery.