'Off-pore' nucleoporins relocalize heterochromatic breaks through phase separation

Abstract

Phase separation forms membraneless compartments in the nuclei, including by establishing heterochromatin "domains" and repair foci. Pericentromeric heterochromatin mostly comprises repeated sequences prone to aberrant recombination, and "safe" homologous recombination (HR) repair of these sequences requires the movement of repair sites to the nuclear periphery before Rad51 recruitment and strand invasion. How this mobilization initiates is unknown, and the contribution of phase separation to these dynamics is unclear. Here, we show that Nup98 nucleoporin is recruited to heterochromatic repair sites before relocalization through Sec13 or Nup88 nucleoporins, and downstream from the Smc5/6 complex and SUMOylation. Remarkably, the phase separation properties of Nup98 are required and sufficient to mobilize repair sites and exclude Rad51, thus preventing aberrant recombination while promoting HR repair. Disrupting this pathway results in heterochromatin repair defects and widespread chromosome rearrangements, revealing a novel "off-pore" role for nucleoporins and phase separation in nuclear dynamics and genome integrity in a multicellular eukaryote. HighlightsO_LINup88 and Sec13 recruit Nup98 to heterochromatic DSBs downstream from Smc5/6 C_LIO_LINup88, Sec13 and Nup98 promote repair focus mobilization in heterochromatin off-pore C_LIO_LINup98 excludes Rad51 from repair sites inside the heterochromatin domain C_LIO_LIPhase separation by Nup98 is required and sufficient for relocalization and Rad51 exclusion C_LI