ATP-driven Rad50 conformations regulate DNA tethering, end resection, and ATM checkpoint signaling

Area of Science:

• Molecular Biology
• Biochemistry
• Genetics

Background:

• The Mre11-Rad50 complex is vital for DNA double-strand break (DSB) repair.
• Mechanisms of Rad50 ATP-dependent regulation in DSB sensing, processing, and signaling remain unclear.

Purpose of the Study:

• To elucidate the role of Rad50 ATP-driven conformational changes in DSB repair.
• To identify specific Rad50 mutations affecting its ATP-bound state and function.

Main Methods:

• Structure-based mutagenesis of Pyrococcus furiosus Rad50.
• X-ray crystallography and X-ray scattering.
• Biochemical assays and functional analyses in model organisms (yeast, S. pombe) and in vitro.

Main Results:

• Identified Rad50 mutants that stabilize or destabilize the ATP-bound state.
• ATP-bound conformation facilitates DNA end binding and tethering.
• ATP hydrolysis-induced opening is essential for DNA resection.
• Altered ATP-bound state stability impairs DNA repair, checkpoint signaling, and ATM activation across species.

Conclusions:

• ATP-dependent Rad50 conformations act as molecular switches.
• These switches regulate Mre11-Rad50 complex functions including DNA tethering, ATM signaling, and resection.
• Provides key insights into the regulation of DNA double-strand break responses.