Open AccessStructure of the human RAD17–RFC clamp loader and 9–1–1 checkpoint clamp bound to a dsDNA–ssDNA junction
Author(s) -
Matthew Day,
Antony W. Oliver,
Laurence H. Pearl
Publication year - 2022
Publication title -
nucleic acids research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 9.008
H-Index - 537
eISSN - 1362-4954
pISSN - 0305-1048
DOI - 10.1093/nar/gkac588
Subject(s) - clamp , biology , dna polymerase delta , dna clamp , microbiology and biotechnology , g2 m dna damage checkpoint , dna , biophysics , dna replication , processivity , dna repair , proliferating cell nuclear antigen , cell cycle checkpoint , cell cycle , biochemistry , rna , reverse transcriptase , cell , mechanical engineering , clamping , engineering , gene
The RAD9-RAD1-HUS1 (9-1-1) clamp forms one half of the DNA damage checkpoint system that signals the presence of substantial regions of single-stranded DNA arising from replication fork collapse or resection of DNA double strand breaks. Loaded at the 5'-recessed end of a dsDNA-ssDNA junction by the RAD17-RFC clamp loader complex, the phosphorylated C-terminal tail of the RAD9 subunit of 9-1-1 engages with the mediator scaffold TOPBP1 which in turn activates the ATR kinase, localised through the interaction of its constitutive partner ATRIP with RPA-coated ssDNA. Using cryogenic electron microscopy (cryoEM) we have determined the structure of a complex of the human RAD17-RFC clamp loader bound to human 9-1-1, engaged with a dsDNA-ssDNA junction. The structure answers the key questions of how RAD17 confers specificity for 9-1-1 over PCNA, and how the clamp loader specifically recognises the recessed 5' DNA end and fixes the orientation of 9-1-1 on the ssDNA.