Related Experiment Video
Updated: Jul 19, 2025
08:53
Strand-Specific Analysis of Proteins at Replicating DNA Strands by Enrichment and Sequencing of Protein-Associated Nascent DNA Method
Published on: May 2, 2025
411
In silico protein interaction screening uncovers DONSON's role in replication initiation
Yang Lim1, Lukas Tamayo-Orrego2, Ernst Schmid1
1Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Blavatnik Institute, Boston, MA 02115, USA.
Science (New York, N.Y.)
|August 17, 2023
Summary
DONSON protein is crucial for eukaryotic DNA replication initiation by assembling the CDC45-MCM2-7-GINS (CMG) helicase. Mutations in DONSON cause microcephalic dwarfism, highlighting its role in human development.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- Eukaryotic DNA replication initiation relies on the assembly of the CDC45-MCM2-7-GINS (CMG) helicase.
- In yeast, a pre-loading complex (pre-LC) facilitates GINS binding to chromatin-associated MCM2-7 to form CMG.
- The precise mechanism of CMG assembly in vertebrates remains incompletely understood.
Purpose of the Study:
- To investigate the role of the metazoan protein DONSON in CMG helicase assembly in vertebrates.
- To elucidate the function of DONSON in the context of replication initiation and its potential link to microcephalic primordial dwarfism.
Main Methods:
- Utilized AlphaFold for in silico screening of protein-protein interactions.
- Performed experimental validation of predicted protein interactions.
- Investigated the impact of a patient-derived DONSON mutation on CMG assembly and its phenotypic consequences in a mouse model.
Main Results:
- DONSON acts as a scaffold for a vertebrate pre-LC, including GINS, TOPBP1, and DNA polymerase epsilon.
- DONSON facilitates the docking of the pre-LC onto MCM2-7, thereby delivering GINS for CMG formation.
- A patient-derived DONSON mutation impairs CMG assembly and causes microcephalic dwarfism in mice.
Conclusions:
- DONSON is essential for CMG helicase assembly in vertebrates, unifying understanding of eukaryotic replication initiation.
- Defective CMG assembly due to DONSON mutations is implicated in microcephalic dwarfism.
- In silico protein-protein interaction screening, exemplified by AlphaFold, accelerates mechanistic biological discovery.
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