The human Rad9/Rad1/Hus1 damage sensor clamp interacts with DNA polymerase beta and increases its DNA substrate utilisation efficiency: implications for DNA repair.
In eukaryotic cells, checkpoints are activated in response to DNA damage. This requires the action of DNA damage sensors such as the Rad family proteins. The three human proteins Rad9, Rad1 and Hus1 form a heterotrimeric complex (called the 9-1-1 complex) that is recruited onto DNA upon damage. DNA damage also triggers the recruitment of DNA repair proteins at the lesion, including specialized DNA polymerases. In this work, we showed that the 9-1-1 complex can physically interact with DNA polymerase beta in vitro. Functional analysis revealed that the 9-1-1 complex had a stimulatory effect on DNA polymerase beta activity. However, the presence of 9-1-1 complex neither affected DNA polymerase lambda, another X family DNA polymerase, nor the two replicative DNA polymerases alpha and delta. DNA polymerase beta stimulation resulted from an increase in its affinity for the primer-template and the interaction with the 9-1-1 complex stimulated deoxyribonucleotides misincorporation by DNA polymerase beta. In addition, the 9-1-1 complex enhanced DNA strand displacement synthesis by DNA polymerase beta on a 1 nt gap DNA substrate. Our data raise the possibility that the 9-1-1 complex might attract DNA polymerase beta to DNA damage sites, thus connecting directly checkpoints and DNA repair.
Institute of Veterinary Biochemistry and Molecular Biology, University of Zürich-Irchel, Wintherturerstrasse 190, CH-8057, Zürich, Switzerland.
Magali
M
Toueille
Nazim
N
El-Andaloussi
Isabelle
I
Frouin
Raimundo
R
Freire
Dorothee
D
Funk
Igor
I
Shevelev
Erica
E
Friedrich-Heineken
Giuseppe
G
Villani
Michael O
MO
Hottiger
Ulrich
U
Hübscher
D001483:Base Sequence; D018797:Cell Cycle Proteins / Q000378:metabolism / Q000502:physiology*; D004247:DNA / Q000378:metabolism*; D004249:DNA Damage; D019951:DNA Polymerase beta / Q000378:metabolism*; D004260:DNA Repair; D003854:Deoxyribonucleotides / Q000378:metabolism; D005092:Exonucleases / Q000378:metabolism / Q000502:physiology*; D006801:Humans; D046911:Macromolecular Substances; D029702:Schizosaccharomyces pombe Proteins; D013698:Templates, Genetic
D018797:Cell Cycle Proteins; D003854:Deoxyribonucleotides; D046911:Macromolecular Substances; D029702:Schizosaccharomyces pombe Proteins; C106332:hus1 protein, S pombe; C069930:rad9 protein; D004247:DNA; D019951:DNA Polymerase beta; D005092:Exonucleases; C113574:Rad1 protein, human