Abstract

Structure-based Engineering for the Advancement of Transposon Tools: The Case of Sleeping Beauty

Franka Voigt, Postdoctoral Fellow, Friedrich Miescher Institute for Biomedical Research, Basel

Transposons are mobile genetic elements with the distinctive ability to autonomously move from one genomic location to another. They constitute a large fraction of modern genomes and their movement (transposition) represents a major force driving genome dynamics and evolution. Moreover, their ability to autonomously move around in genomes allowed their application as powerful genetic engineering tools. The reconstructed Sleeping Beauty (SB) transposon has revolutionized the functional genomics analysis of vertebrates and found widespread applications in transgenesis including human gene therapy. Nevertheless, the mechanism ofSB transposition is poorly understood and no structural data is available, limiting further design of SB based genetic engineering tools. In this talk, I will present our recent biochemical and structural results that provide long sought insights into SB transposition. Using biochemistry and cell biology approaches, we have described the first steps of SB transposition and show that they follow a distinct pathway, different from homologous transposons. We have also determined the first crystal structure of the SB transposase catalytic domain, which provides important insights into the mechanism of target DNA recognition and integration. In addition, the structure explains hyperactive mutations in the SB transposase and allows us to engineer further improved variants.