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

Area of Science:

• Cell biology
• Molecular genetics
• Genomics

Background:

• Membraneless compartments form via phase separation in the nucleus, organizing heterochromatin and DNA repair foci.
• Pericentromeric heterochromatin contains repetitive sequences susceptible to aberrant recombination.
• Safe homologous recombination (HR) repair of these sequences necessitates movement to the nuclear periphery prior to Rad51 recruitment.

Purpose of the Study:

• To investigate the mechanisms initiating the mobilization of heterochromatic repair sites.
• To elucidate the role of phase separation in these nuclear dynamics.
• To understand the contribution of nucleoporins to heterochromatin repair and genome stability.

Main Methods:

• Recruitment assays for Nup98 at heterochromatic repair sites.
• Analysis of nucleoporin relocalization dynamics (Sec13, Nup88).
• Investigating the role of Smc5/6 complex and SUMOylation.
• Assessing the impact of Nup98 phase separation properties on repair site mobilization and Rad51 exclusion.
• Evaluating heterochromatin repair and chromosome integrity upon pathway disruption.

Main Results:

• Nup98 nucleoporin is recruited to heterochromatic repair sites before relocalization via Sec13 or Nup88.
• Nup98 recruitment occurs downstream of the Smc5/6 complex and SUMOylation.
• Nup98's phase separation properties are essential and sufficient for mobilizing repair sites and excluding Rad51.
• Disruption of this pathway leads to heterochromatin repair defects and widespread chromosome rearrangements.

Conclusions:

• Nup98-mediated phase separation drives the mobilization of DNA repair sites to the nuclear periphery.
• This process is crucial for preventing aberrant recombination and ensuring faithful HR repair in heterochromatin.
• Nucleoporins play a novel "off-pore" role in nuclear dynamics and maintaining genome integrity through phase separation.