Molecular mechanisms of integration site selection by Dictyostelium retrotransposons

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Parasitism of genomes by mobile elements is a general phenomenon in biology. Although it is widely accepted that the activity of mobile elements may be beneficial for genome evolution of their hosts, there is also evidence that excessive amplification of selfish mobile elements may reduce host fitness by compromising genome stability. Thus, the study of interaction of mobile elements with their hosts genomes allows fundamental insights into genome evolution, the basis of organismal evolution.Especially in genomes with high gene density and reduced intergenic regions, mobile elements must evolve strategies to avoid insertional mutagenesis of host genes. Retrotransposons in the yeast Saccharomyces cerevisiae and the social amoeba Dictyostelium discoideum are intriguing examples of convergent evolution to solve this problem by targeting to the flanking regions of tRNA genes. Another means to avoid insertional mutagenesis of host genes, which is observed in genomes of plants, fungi and dictyostelid amoebae, seems to be the accumulation of mobile elements in gene-poor regions in heterochromatin and centromeric DNA. The D. discoideum genome accommodates retrotransposons of different phylogenetic origin that accumulate either in euchromatin in tRNA gene-flanking regions and in centromeric heterochromatin. Therefore, D. discoideum is an exceptional model to study different mechanisms of targeted integration by mobile elements and results obtained in this project will further expand the general knowledge about how dynamic parasite-host interactions contribute to genome evolution. In the first set of experiments we focus on retrotransposon TRE5-A. This element integrates with high precision upstream of tRNA genes in a presumed nucleosome-free region. Because TRE5-A integration depends on the host factor CbfA that contains a JmjC domain, a putative histone demethylase, we determine whether a CbfA-mediated nucleosome remodeling activity upstream of tRNA genes is required for position-specific integration of TRE5-A. In addition, we determine the RNA polymerase III transcription complex distribution in the D. discoideum genome to obtain direct proof that interaction with such complexes determines TRE5-A integration site selection. In the second set of experiments we compare the related DGLT-A and Skipper elements to evaluate what kind of interaction with chromatin determines integration at tRNA genes (DGLT-A) or in heterochromatin (Skipper). We extend preliminary data and determine whether interaction of the DGLT-A ribonuclease H domain with TFIIIC subunit Tfc4 determines integration site selection at tRNA genes in vivo. Further, we evaluate whether chromo domains in Skipper-like elements allow chromoviruses to integrate in heterochromatin, and whether functional degeneration of chromo domains may allow such elements to evolve new targeting strategies to colonize euchromatic regions of their hosts genomes.