Spatial and temporal organization of the Bacillus subtilis replication cycle

Mol Microbiol. 2006 Oct;62(1):57-71. doi: 10.1111/j.1365-2958.2006.05356.x. Epub 2006 Aug 30.

Abstract

DNA replication occurs at discrete sites in the cell. To gain insight into the spatial and temporal organization of the Bacillus subtilis replication cycle, we simultaneously visualized replication origins and the replication machinery (replisomes) inside live cells. We found that the origin of replication is positioned near midcell prior to replication. After initiation, the replisome colocalizes with the origin, confirming that replication initiates near midcell. The replisome remains near midcell after duplicated origins separate. Artificially mispositioning the origin region leads to mislocalization of the replisome indicating that the location of the origin at the time of initiation establishes the position of the replisome. Time-lapse microscopy revealed that a single replisome focus reversibly splits into two closely spaced foci every few seconds in many cells, including cells that recently initiated replication. Thus, sister replication forks are likely not intimately associated with each other throughout the replication cycle. Fork dynamics persisted when replication elongation was halted, and is thus independent of the relative movement of DNA through the replisome. Our results provide new insights into how the replisome is positioned in the cell and refine our current understanding of the spatial and temporal events of the B. subtilis replication cycle.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacillus subtilis / cytology
  • Bacillus subtilis / genetics*
  • Bacillus subtilis / metabolism
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism
  • Bacterial Proteins / physiology
  • Cell Cycle / genetics
  • Cell Cycle / physiology
  • Cell Division / genetics
  • Cell Division / physiology
  • Chromosomes, Bacterial / genetics
  • DNA Replication / genetics*
  • DNA Replication / physiology
  • Gene Expression Regulation, Bacterial / genetics
  • Green Fluorescent Proteins
  • Luminescent Proteins
  • Microscopy, Fluorescence
  • Models, Biological
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Replication Origin / genetics
  • Replication Origin / physiology

Substances

  • Bacterial Proteins
  • Luminescent Proteins
  • Recombinant Fusion Proteins
  • yellow fluorescent protein, Bacteria
  • Green Fluorescent Proteins