Recruitment of Fkh1 to replication origins requires precisely positioned Fkh1/2 binding sites and concurrent assembly of the pre-replicative complex

Autor: Nikita Avvakumov, Kristiina Jalakas, Allan Reinapae, Arnold Kristjuhan, Marko Lõoke, Kersti Kristjuhan
Jazyk: angličtina
Rok vydání: 2017
Předmět:
0301 basic medicine
Cancer Research
Synthesis Phase
Cell Cycle Proteins
Yeast and Fungal Models
Biochemistry
S Phase
Database and Informatics Methods
Cell Cycle and Cell Division
Genetics (clinical)
Genetics
Insertion Mutation
Forkhead Transcription Factors
Cell biology
Chromatin
Nucleic acids
Experimental Organism Systems
Cell Processes
Sequence Analysis
Protein Binding
Research Article
DNA re-replication
Pre-replicative complex
Saccharomyces cerevisiae Proteins
lcsh:QH426-470
Bioinformatics
Replication Origin
Saccharomyces cerevisiae
Biology
DNA replication
Research and Analysis Methods
03 medical and health sciences
Saccharomyces
Model Organisms
Control of chromosome duplication
Sequence Motif Analysis
DNA-binding proteins
Molecular Biology
Ecology
Evolution
Behavior and Systematics
Binding Sites
Biology and life sciences
Organisms
Fungi
Proteins
DNA
Cell Biology
Yeast
DNA binding site
lcsh:Genetics
030104 developmental biology
Genetic Loci
Mutation
Origin recognition complex
Zdroj: PLoS Genetics, Vol 13, Iss 1, p e1006588 (2017)
PLoS Genetics
ISSN: 1553-7404
1553-7390
Popis: In budding yeast, activation of many DNA replication origins is regulated by their chromatin environment, whereas others fire in early S phase regardless of their chromosomal location. Several location-independent origins contain at least two divergently oriented binding sites for Forkhead (Fkh) transcription factors in close proximity to their ARS consensus sequence. To explore whether recruitment of Forkhead proteins to replication origins is dependent on the spatial arrangement of Fkh1/2 binding sites, we changed the spacing and orientation of the sites in early replication origins ARS305 and ARS607. We followed recruitment of the Fkh1 protein to origins by chromatin immunoprecipitation and tested the ability of these origins to fire in early S phase. Our results demonstrate that precise spatial and directional arrangement of Fkh1/2 sites is crucial for efficient binding of the Fkh1 protein and for early firing of the origins. We also show that recruitment of Fkh1 to the origins depends on formation of the pre-replicative complex (pre-RC) and loading of the Mcm2-7 helicase, indicating that the origins are regulated by cooperative action of Fkh1 and the pre-RC. These results reveal that DNA binding of Forkhead factors does not depend merely on the presence of its binding sites but on their precise arrangement and is strongly influenced by other protein complexes in the vicinity.
Author summary In this study, we explore the mechanisms that determine activation of DNA replication origins in early S phase. It has been shown that a subset of replication origins is regulated by Forkhead family transcription factors that ensure their firing at the beginning of S phase. However, the recruitment of Forkhead factors to replication origins is not a straightforward process–there are thousands of Forkhead binding sites in the genome and their presence does not guarantee that Forkheads actually bind these sites. We show that recruitment of Fkh1 protein to DNA replication origins requires precise arrangement of Forkhead binding sites and depends on formation of pre-replicative complexes at the origins. These results clarify the mechanisms of Forkhead-dependent regulation of early DNA replication origins and also reveal that mere presence of consensus binding sites is not sufficient for recruitment of Forkhead proteins to their target loci.
Databáze: OpenAIRE
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