Physical Interaction between Human RAD52 and RPA Is Required for Homologous Recombination in Mammalian Cells

Min S Park,Dale L Ludwig,Evelyn Stigger,Suk-Hee Lee

doi:10.1074/jbc.271.31.18996

Abstract

The yeast RAD52 protein is essential for DNA double-strand break repair, and meiotic and mitotic recombination. RPA is a protein complex of three subunits (70, 34, and 11 kDa) that has been shown to be involved in DNA replication, nucleotide excision repair, and homologous recombination. Here, we demonstrate a physical interaction between human RAD52 and RPA in vivo and in vitro. In addition, the domain (amino acids 221-280) in RAD52 protein that mediates the interaction with the 34-kDa subunit of RPA was also determined. Overexpression of mutant RAD52 proteins lacking the interaction domain (amino acids 221-240, 241-260, and 261-280) failed to induce homologous recombination in monkey cells. We have previously shown that overexpression of human RAD52 induced homologous recombination in these cells. These results suggest that direct physical interactions between RAD52 and RPA are essential for homologous recombination in mammalian cells.

Highlights

The yeast RAD52 protein is essential for DNA doublestrand break repair, and meiotic and mitotic recombination
These results suggest that direct physical interactions between RAD52 and RPA are essential for homologous recombination in mammalian cells
We have recently shown that RPA-XPA interactions can lead to inhibition of replication, which may suggest that DNA repair overrides DNA replication in vitro through a direct interaction between RPA and XPA [20]

Summary

Introduction

The yeast RAD52 protein is essential for DNA doublestrand break repair, and meiotic and mitotic recombination. We have previously shown that overexpression of human RAD52 induced homologous recombination in these cells These results suggest that direct physical interactions between RAD52 and RPA are essential for homologous recombination in mammalian cells. Most of the information regarding the functions of the RAD52 protein has come from genetic studies in yeast These studies suggested that the RAD52 protein is not required for the initiation of recombination, but is essential for the intermediate stage following the formation of double strand breaks but before the appearance of stable recombinants [4]. Human RAD52 protein has been shown to confer resistance to ionizing radiation and induce homologous recombination in monkey cells [5] This indicates that the RAD52mediated homologous recombination plays a role in double strand break repair in mammalian cells. We demonstrated the physical interaction between human RAD52 and RPA and addressed the importance of this intermolecular interaction in homologous recombination in monkey cells

Methods

Results

Conclusion