Sld3 is essential for the initiation of DNA replication, but Sld3 does not travel with a replication fork. GINS binds to Cdc45 and Mcm2-7 to form the replication fork helicase in eukaryotes. We purified Sld3, Cdc45, GINS, and Mcm2-7 and studied their interaction and assembly into complexes. Sld3 binds tightly to Cdc45 in the presence or absence of cyclin-dependent kinase activity. Furthermore, Sld3 binds tightly to the Mcm2-7 complex, and a ternary complex forms among Cdc45, Mcm2-7, and Sld3, with a 1:1:1 stoichiometry (CMS complex). GINS binds directly to Mcm2-7, and GINS competes with Sld3 for Mcm2-7 binding. GINS also binds directly to Cdc45, and GINS competes with Sld3 for Cdc45 binding. Cdc45, Mcm2-7, and GINS form a ternary complex with a stoichiometry of 1:1:1 (CMG complex). Size exclusion data reveal that when Sld3, Cdc45, Mcm2-7, and GINS are added together, the result is a mixture of CMS and CMG complexes. The data suggest that GINS and Sld3 compete with one another for Mcm2-7 and Cdc45 binding. Our results are consistent with a model wherein GINS trades places with Sld3 at a replication origin, contributing to the activation of the replication fork helicase.
Download full-text PDF |
Source |
|---|---|
| http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3077617 | PMC |
| http://dx.doi.org/10.1074/jbc.M111.218305 | DOI Listing |
Publication Analysis
Top Keywords
Similar Publications
Faithful genome duplication is essential for preserving the genetic stability of dividing cells. DNA replication is carried out during the S phase by a dynamic complex of proteins termed the replisome. At the heart of the replisome is the CDC45-MCM2-7-GINS (CMG) helicase, which separates the two strands of the DNA double helix such that DNA polymerases can copy each strand.
View Article and Find Full Text PDFUSP37 prevents unscheduled replisome unloading through MCM complex deubiquitination.
bioRxiv
September 2024
Department of Pharmacology and Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, NC 27599, USA.
The CMG helicase (CDC45-MCM2-7-GINS) unwinds DNA as a component of eukaryotic replisomes. Replisome (dis)assembly is tightly coordinated with cell cycle progression to ensure genome stability. However, factors that prevent premature CMG unloading and replisome disassembly are poorly described.
View Article and Find Full Text PDFUSP37 prevents premature disassembly of stressed replisomes by TRAIP.
bioRxiv
September 2024
Cancer Research UK Cambridge Institute, Li Ka Shing Building, Robinson Way, Cambridge CB2 0RE, UK.
The E3 ubiquitin ligase TRAIP associates with the replisome and helps this molecular machine deal with replication stress. Thus, TRAIP promotes DNA inter-strand crosslink repair by triggering the disassembly of CDC45-MCM2-7-GINS (CMG) helicases that have converged on these lesions. However, disassembly of single CMGs that have stalled temporarily would be deleterious, suggesting that TRAIP must be carefully regulated.
View Article and Find Full Text PDFAssembly, Activation, and Helicase Actions of MCM2-7: Transition from Inactive MCM2-7 Double Hexamers to Active Replication Forks.
Biology (Basel)
August 2024
Genome Dynamics Project, Department of Basic Medical Sciences, Tokyo Metropolitan Institute of Medical Science, Setagaya-ku, Tokyo 156-8506, Japan.
In this review, we summarize the processes of the assembly of multi-protein replisomes at the origins of replication. Replication licensing, the loading of inactive minichromosome maintenance double hexamers (dhMCM2-7) during the G1 phase, is followed by origin firing triggered by two serine-threonine kinases, Cdc7 (DDK) and CDK, leading to the assembly and activation of Cdc45/MCM2-7/GINS (CMG) helicases at the entry into the S phase and the formation of replisomes for bidirectional DNA synthesis. Biochemical and structural analyses of the recruitment of initiation or firing factors to the dhMCM2-7 for the formation of an active helicase and those of origin melting and DNA unwinding support the steric exclusion unwinding model of the CMG helicase.
View Article and Find Full Text PDFReplisome Proximal Protein Associations and Dynamic Proteomic Changes at Stalled Replication Forks.
Mol Cell Proteomics
May 2024
Faculty of Medicine and Health Sciences, Department of Immunology and Cell Biology, Université de Sherbrooke, Sherbrooke, Québec, Canada. Electronic address:
Article Synopsis
- - DNA replication is essential for cell division, involving precise coordination of various proteins at replication forks during the S phase, and is sensitive to genotoxic stressors, which can disrupt the process and lead to the need for DNA repair mechanisms.
- - Researchers used proximity-dependent biotin identification to explore interactions among 17 proteins related to core replication components, revealing a complex network of protein associations in the context of stress induced by hydroxyurea.
- - The study found that hydroxyurea treatment altered the relationships among proteins involved in DNA replication, identifying new potential players in the cell's response to replication challenges, and establishing a foundational framework for understanding cellular reactions to DNA replication issues.
Want AI Summaries of new PubMed Abstracts delivered to your In-box?
Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!