A hand-off mechanism for primosome assembly in replication restart.
Collapsed DNA replication forks must be reactivated through origin-independent reloading of the replication machinery (replisome) to ensure complete duplication of cellular genomes. In E. coli, the PriA-dependent pathway is the major replication restart mechanism and requires primosome proteins PriA, PriB, and DnaT for replisome reloading. However, the molecular mechanisms that regulate origin-independent replisome loading are not fully understood. Here, we demonstrate that assembly of primosome protein complexes represents a key regulatory mechanism, as inherently weak PriA-PriB and PriB-DnaT interactions are strongly stimulated by single-stranded DNA. Furthermore, the binding site on PriB for single-stranded DNA partially overlaps the binding sites for PriA and DnaT, suggesting a dynamic primosome assembly process in which single-stranded DNA is handed off from one primosome protein to another as a repaired replication fork is reactivated. This model helps explain how origin-independent initiation of DNA replication is restricted to repaired replication forks, preventing overreplication of the genome.
Lopper
Matthew
M
Department of Biomolecular Chemistry, University of Wisconsin School of Medicine and Public Health, 550 Medical Sciences Center, 1300 University Avenue, Madison, WI 53706, USA.
Boonsombat
Ruethairat
R
Sandler
Steven J
SJ
Keck
James L
JL
D001665:Binding Sites / Q000502:physiology; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism*; D004261:DNA Replication / Q000502:physiology*; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004259:DNA-Directed DNA Polymerase / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D016680:Genome, Bacterial / Q000502:physiology; D008954:Models, Biological; D009097:Multienzyme Complexes / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding / Q000502:physiology; D018741:Replication Origin; D000099272:Replisomes
D004265:DNA Helicases; D004269:DNA, Bacterial; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D004259:DNA-Directed DNA Polymerase; D029968:Escherichia coli Proteins; D009097:Multienzyme Complexes; D000099272:Replisomes; C104198:DnaT protein, E coli; C086004:PriB protein, E coli; C506743:priA protein, E coli