Understanding the in vivo functions of Holliday junction resolvase Yen1

Elaborate replication fork recovery pathways support the duplication of the genome under replication stress. Nucleases and helicases take center stage in these pathways, but our understanding of the molecular underpinnings remains incomplete. The Holliday junction resolving nucleases Mus81-Mms4 (human MUS81-EME1) and Yen1 (GEN1) act redundantly to remove replication-associated homologous recombination repair intermediates, safeguarding chromosome segregation. They are cell cycle regulated, so that Mus81-Mms4/MUS81-EME1 activity peaks first, making it the major resolvase in yeast and human. Yen1/GEN1 subsequently provides a catchall activity for the resolution of recombinational joint molecules that escape the attention of Mus81-Mms4.
In this work, we used budding yeast as a model to explore whether additional, Mus81-independent functions of Yen1 exist, potentially outside the context of canonical Holliday junction resolution. To this end, we investigated the reported genetic interactome of YEN1. We disproved a purported genetic interaction between YEN1 and PBY1, linking wrongly assigned PBY1 interactions to MUS81-MMS4 instead. More significantly, addressing a synthetic sick interaction between YEN1 and the conserved nuclease-helicase DNA2, we identify a first unique function of Yen1 in targeting replication, as opposed to recombination, intermediates. Furthermore, we uncover a novel role for the elusive Dna2 helicase activity in the recovery of stalled replication intermediates. Our findings provide new insight into the replication stress response in eukaryotes.