Xaver, M.; Shodhan, A.; Kashko, N.; Lichten, M.; Matos, J.

Holliday junctions (HJs) are four-way branched DNA intermediates in double strand break (DSB) repair by homologous recombination. During meiosis, HJs are resolved predominantly as crossovers that are necessary for correct chromosome segregation. However, the genome-wide behavior of meiotic HJs has remained inaccessible without direct mapping. By a HJ-binding peptide approach (HJSeq), we capture the genome-wide distribution of these crucial intermediates for the first time. We show that during pachytene, the stage before crossover formation, HJs undergo sustained directional branch migration, reorganizing the HJ-landscape in a transcription-linked manner, away from DSB sites towards convergent transcription sites. The presented findings establish pachytene as an active remodeling phase in which HJ branch-migration accommodates an active chromatin environment to support regulated crossover formation and faithful chromosome segregation.