Crystal Structure of the Rad9-Rad1-Hus1 DNA Damage Checkpoint Complex¿Implications for Clamp Loading and Regulation

Rad9, Rad1, and Hus1 form a heterotrimeric complex (9-1-1) that is loaded onto DNA at sites of DNA damage. DNA-loaded 9-1-1 activates signaling through the Chk1 arm of the DNA damage checkpoint response via recruitment and stimulation of ATR. Additionally, 9-1-1 may play a direct role in facilitating DNA damage repair via interaction with a number of DNA repair enzymes. We have now determined the crystal structure of the human 9-1-1 complex, revealing a toroidal structure with a similar architecture to the homotrimeric PCNA DNA-binding clamp. The structure explains the formation of a unique heterotrimeric arrangement and reveals significant differences among the three subunits in the sites implicated in binding to the clamp loader and to ligand proteins. Biochemical analysis reveals a single repair enzyme-binding site on 9-1-1 that can be blocked competitively by the PCNA-binding cell-cycle regulator p21