DNA double-strand break repair pathway choice is directed by distinct MRE11 nuclease activities

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• The MRE11-RAD50-NBS1 (MRN) complex, particularly MRE11, is crucial for DNA double-strand break repair (DSBR), detection, and signaling.
• The precise roles of MRE11's endonuclease and exonuclease activities in directing nonhomologous end-joining (NHEJ) versus homologous recombination (HR) remain unclear.

Purpose of the Study:

• To discover specific MRE11 endonuclease or exonuclease inhibitors using structure-based design and a focused chemical library.
• To investigate how these inhibitors influence DNA repair pathway choice at double-strand breaks (DSBs) in G2-phase cells after radiation exposure.

Main Methods:

• Structure-based drug design was used to create a focused chemical library targeting MRE11 nuclease activities.
• Experiments involved generating DSBs in G2-phase cells via radiation and assessing repair pathway choice using specific MRE11 inhibitors.
• Replication protein A (RPA) chromatin binding was monitored to evaluate DNA resection status.

Main Results:

• Inhibition of MRE11 nucleases impaired radiation-induced RPA chromatin binding, indicating reduced DNA resection.
• MRE11 endonuclease inhibition promoted NHEJ over HR, while MRE11 exonuclease inhibition resulted in a significant repair defect.
• Distinct roles for MRE11's endonuclease and exonuclease activities in regulating DSBR pathway choice were defined.

Conclusions:

• Nuclease-specific MRE11 inhibitors were successfully developed, providing tools to dissect DSBR mechanisms.
• MRE11 endonuclease activity is proposed to initiate DNA resection, licensing HR.
• MRE11 exonuclease activity, along with EXO1/BLM, is implicated in subsequent bidirectional resection, committing the cell to HR.