Cdc6-induced conformational changes in ORC bound to origin DNA revealed by cryo-electron microscopy

Area of Science:

• Molecular Biology
• Structural Biology
• Genetics

Background:

• The eukaryotic origin recognition complex (ORC) is crucial for DNA replication initiation.
• ORC interacts with replication origins to remodel DNA before S phase.
• Understanding ORC's structure and DNA interaction is key to replication control.

Purpose of the Study:

• To determine the structure of the Saccharomyces cerevisiae ORC-Cdc6-DNA complex.
• To elucidate the mechanism of origin DNA binding and remodeling by ORC and Cdc6.
• To provide insights into the initiation of eukaryotic DNA replication.

Main Methods:

• Single-particle cryo-electron microscopy (cryo-EM) was used to determine the structure.
• The study analyzed the supramolecular assembly of ORC, Cdc6, and ARS1 origin DNA.
• Computational methods were used for segmentation and docking with homologous structures.

Main Results:

• A high-resolution cryo-EM structure of the ORC-Cdc6-ARS1 DNA complex was obtained.
• The six ORC subunits (Orc1-Orc6) and Cdc6 were visualized in complex with DNA.
• ORC undergoes conformational changes upon Cdc6 binding, reorienting the Orc1 N-terminal BAH domain.
• A model for DNA binding suggests ORC bends and wraps the origin DNA along its interior surface.
• Cdc6 binding extends the ORC footprint on origin DNA.

Conclusions:

• The study reveals a detailed structural mechanism for origin recognition and DNA remodeling.
• ORC-Cdc6 interaction is critical for preparing origins for replication initiation.
• The findings advance our understanding of the fundamental process of DNA replication in eukaryotes.