3D-SIM Super Resolution Microscopy Reveals a Bead-Like Arrangement for FtsZ and the Division Machinery: Implications for Triggering Cytokinesis

PLoS Biol. 2012;10(9):e1001389. doi: 10.1371/journal.pbio.1001389. Epub 2012 Sep 11.

Abstract

FtsZ is a tubulin-like GTPase that is the major cytoskeletal protein in bacterial cell division. It polymerizes into a ring, called the Z ring, at the division site and acts as a scaffold to recruit other division proteins to this site as well as providing a contractile force for cytokinesis. To understand how FtsZ performs these functions, the in vivo architecture of the Z ring needs to be established, as well as how this structure constricts to enable cytokinesis. Conventional wide-field fluorescence microscopy depicts the Z ring as a continuous structure of uniform density. Here we use a form of super resolution microscopy, known as 3D-structured illumination microscopy (3D-SIM), to examine the architecture of the Z ring in cells of two Gram-positive organisms that have different cell shapes: the rod-shaped Bacillus subtilis and the coccoid Staphylococcus aureus. We show that in both organisms the Z ring is composed of a heterogeneous distribution of FtsZ. In addition, gaps of fluorescence were evident, which suggest that it is a discontinuous structure. Time-lapse studies using an advanced form of fast live 3D-SIM (Blaze) support a model of FtsZ localization within the Z ring that is dynamic and remains distributed in a heterogeneous manner. However, FtsZ dynamics alone do not trigger the constriction of the Z ring to allow cytokinesis. Lastly, we visualize other components of the divisome and show that they also adopt a bead-like localization pattern at the future division site. Our data lead us to propose that FtsZ guides the divisome to adopt a similar localization pattern to ensure Z ring constriction only proceeds following the assembly of a mature divisome.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacillus subtilis / cytology*
  • Bacillus subtilis / metabolism
  • Bacterial Proteins / metabolism*
  • Cytokinesis*
  • Cytoskeletal Proteins / metabolism*
  • Green Fluorescent Proteins / metabolism
  • Imaging, Three-Dimensional / methods*
  • Microbial Viability
  • Microscopy / methods*
  • Models, Biological
  • Movement
  • Protein Transport
  • Recombinant Fusion Proteins / metabolism
  • Staphylococcus aureus / cytology*
  • Staphylococcus aureus / metabolism
  • Time-Lapse Imaging

Substances

  • Bacterial Proteins
  • Cytoskeletal Proteins
  • FtsZ protein, Bacteria
  • Recombinant Fusion Proteins
  • Green Fluorescent Proteins

Grant support

This work was funded by an ARC Discovery grant DP0666670 to EJH. CBW was funded by an Australian National Health and Medical Research Council Senior Research Fellowship. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.