Rad53 regulates the lifetime of Rdh54 at homologous recombination intermediates

Abstract

Rdh54 is a conserved DNA translocase that participates in homologous recombination (HR), DNA checkpoint adaptation, and chromosome segregation. Saccharomyces cerevisiae Rdh54 is a known target of the Mec1/Rad53 signaling axis, which globally protects genome integrity during DNA metabolism. While phosphorylation of DNA repair proteins by Mec1/Rad53 is critical for HR progression little is known about how specific post translational modifications alter HR reactions. Phosphorylation of Rdh54 is linked to protection of genomic integrity but the consequences of modification remain poorly understood. Here, we demonstrate that phosphorylation of the Rdh54 C-terminus by the effector kinase Rad53 down regulates Rdh54 activity resulting in a 50% decrease in translocation velocity on dsDNA as revealed by single molecule imaging. Genetic assays reveal that loss of phosphorylation reduces interhomolog gene conversion, increases break induced replication, and decreases crossover outcomes. Our data highlight Rad53 as a key regulator of HR intermediates through activation and attenuation of Rdh54 motor function.