ATM/ATR-mediated phosphorylation of PALB2 promotes RAD51 function

Area of Science:

• Molecular Biology
• Cellular Biology
• Genetics

Background:

• DNA damage activates ATM and ATR kinases, coordinating cell cycle checkpoints and DNA repair.
• Homology-directed repair (HDR) relies on RAD51 nucleofilament formation, a process involving PALB2-BRCA2.
• The specific roles of ATM and ATR in RAD51 nucleofilament formation during HDR are not well understood.

Purpose of the Study:

• To investigate the roles of ATM and ATR kinases in the regulation of PALB2 during DNA repair.
• To determine the impact of PALB2 phosphorylation on RAD51 foci formation and HDR efficiency.
• To elucidate the functional significance of PALB2 phosphorylation in response to genotoxic stress.

Main Methods:

• Genotoxic stress induction (ionizing radiation, hydroxyurea).
• Phosphorylation site analysis of PALB2 using phospho-deficient and phospho-mimicking mutants.
• Assessment of RAD51 foci formation and HDR activity in cells expressing PALB2 mutants.
• Evaluation of DNA damage checkpoint response.

Main Results:

• PALB2 is significantly phosphorylated upon genotoxic stress, mediated by ATM and ATR kinases at N-terminal S/Q sites.
• A phospho-deficient PALB2 mutant impairs RAD51 foci formation and reduces HDR efficiency compared to wild-type.
• A phospho-mimicking PALB2 mutant supports RAD51 foci formation.
• The DNA damage checkpoint response remains functional in cells expressing the phospho-deficient PALB2 mutant, indicating functional separation.

Conclusions:

• PALB2 phosphorylation by ATM/ATR is a critical regulatory mechanism in DNA repair.
• This phosphorylation event is essential for proper RAD51 nucleofilament assembly and efficient HDR.
• PALB2 phosphorylation represents a novel regulatory step in maintaining genome stability following DNA damage.