Specific basic patch-dependent multimerization of Saccharomyces cerevisiae ORC on single-stranded DNA promotes ATP
Hironori Kawakami1, Ryuya Muraoka1, Eiji Ohashi2
1Department of Molecular Biology, Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka, Japan.
Summary
The eukaryotic origin sensor (EOS) regulates origin recognition complexes (ORC) on single-stranded DNA, promoting ORC multimerization and ATP hydrolysis. This mechanism may control ORC activity at specific genomic sites.
Area of Science:
- Molecular Biology
- Genetics
- Cell Biology
Background:
- Replication initiation at specific genomic loci is crucial for genetic inheritance.
- Origin Recognition Complexes (ORCs) bind DNA origins to recruit the replicative helicase.
- The eukaryotic origin sensor (EOS) in Orc1 recognizes essential origin DNA regions.
Purpose of the Study:
- To investigate whether the EOS regulates ORC binding to single-stranded DNA (ssDNA).
- To understand the mechanism of ORC-ssDNA interaction and its functional consequences.
Main Methods:
- In vitro biochemical assays to study ORC multimerization on ssDNA.
- Analysis of genomics data to identify in vivo ORC-ssDNA binding sites.
- Assays to measure ORC ATPase activity.
Main Results:
- ORC multimerizes on ssDNA in vitro independently of adenine nucleotides.
- ORC multimerization on ssDNA is dependent on the EOS.
- ORC multimerization stimulates ORC ATPase activity.
- Genomics data suggest in vivo ORC-ssDNA binding occurs outside replication origins.
Conclusions:
- EOS function is differentiated on ssDNA, promoting ORC self-assembly and ATP hydrolysis.
- These mechanisms may modulate ORC activity at specific genomic loci.
- The findings suggest a conserved mechanism among eukaryotes for regulating DNA replication.
Keywords:
AAA+ATPaseDNA replicationintrinsically disordered regionorigin recognition complexprotein multimerizationsingle-stranded DNAMore Related Videos
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