In silico protein interaction screening uncovers DONSON's role in vertebrate CMG helicase assembly

Yang Lim, Lukas Tamayo Orrego, Ernst Schmid, Zygimante Tarnauskaite, Olga V. Kochenova, Rhian Gruar, Sachiko Muramatsu, Luke Lynch, Aitana Verdu Schlie, Paula L Carroll, Gheorghe Chistol, Martin A M Reijns, Masato T. Kanemaki, Andrew P Jackson*, Johannes C. Walter*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

Abstract / Description of output

: CMG (Cdc45-MCM2-7-GINS) helicase assembly is the central event in eukaryotic
replication initiation. In yeast, a multi-subunit “pre-Loading Complex” (pre-LC) accompanies
GINS to chromatin-bound MCM2-7, leading to CMG formation. Here, we report that DONSON,
30 a metazoan protein mutated in microcephalic primordial dwarfism, is required for CMG assembly
in vertebrates. Using AlphaFold to screen for protein-protein interactions followed by
experimental validation, we show that DONSON scaffolds a vertebrate pre-LC containing GINS,
TOPBP1, and DNA pol e. Our evidence suggests that DONSON docks the pre-LC onto MCM2-
7, delivering GINS to its binding site in CMG. A patient-derived DONSON mutation compromises
35 CMG assembly and recapitulates microcephalic dwarfism in mice. These results unify our
understanding of eukaryotic replication initiation, implicate defective CMG assembly in
microcephalic dwarfism, and illustrate how in silico protein-protein interaction screening
accelerates mechanistic discovery
Original languageEnglish
Early online date17 Aug 2023
Publication statusE-pub ahead of print - 17 Aug 2023


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