Massively parallel jumping assay decodes Alu retrotransposition activity

The human genome contains millions of copies of retrotransposons that are silenced but many of these copies have potential to become active if mutated, having phenotypic consequences, including disease. However, it is not thoroughly understood how nucleotide changes in retrotransposons affect their jumping activity. Here, we develop a massively parallel jumping assay (MPJA) that tests the jumping potential of thousands of transposons en masse. We generate a nucleotide variant library of four Alu retrotransposons containing 165,087 different haplotypes and test them for their jumping ability using MPJA. We found 66,821 unique jumping haplotypes, allowing us to pinpoint domains and variants vital for transposition. Mapping these variants to the Alu-RNA secondary structure revealed stem-loop features that contribute to jumping potential. Combined, our work provides a high-throughput assay that assesses the ability of retrotransposons to jump and identifies nucleotide changes that have the potential to reactivate them in the human genome.