Dynamic SpoIIIE assembly mediates septal membrane fission during Bacillus subtilis sporulation.
SpoIIIE is an FtsK-related protein that transports the forespore chromosome across the Bacillus subtilis sporulation septum. We use membrane photobleaching and protoplast assays to demonstrate that SpoIIIE is required for septal membrane fission in the presence of trapped DNA, and that DNA is transported across separate daughter cell membranes, suggesting that SpoIIIE forms a channel that partitions the daughter cell membranes. Our results reveal a close correlation between septal membrane fission and the assembly of a stable SpoIIIE translocation complex at the septal midpoint. Time-lapse epifluorescence, total internal reflection fluorescence (TIRF) microscopy, and live-cell photoactivation localization microscopy (PALM) demonstrate that the SpoIIIE transmembrane domain mediates dynamic localization to active division sites, whereas the assembly of a stable focus also requires the cytoplasmic domain. The transmembrane domain fails to completely separate the membrane, and it assembles unstable foci. TIRF microscopy and biophysical modeling of fluorescence recovery after photobleaching (FRAP) data suggest that this unstable protein transitions between disassembled and assembled oligomeric states. We propose a new model for the role of SpoIIIE assembly in septal membrane fission that has strong implications for how the chromosome terminus crosses the septum.
Division of Biological Sciences, University of California at San Diego, La Jolla, California 92093, USA.
Tinya C
TC
Fleming
Jae Yen
JY
Shin
Sang-Hyuk
SH
Lee
Eric
E
Becker
Kerwyn Casey
KC
Huang
Carlos
C
Bustamante
Kit
K
Pogliano
D001412:Bacillus subtilis / Q000235:genetics / Q000254:growth & development / Q000378:metabolism / Q000502:physiology*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism*; D004271:DNA, Fungal / Q000378:metabolism; D036681:Fluorescence Recovery After Photobleaching; D008954:Models, Biological; D009154:Mutation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D013171:Spores, Bacterial / Q000254:growth & development*
D001426:Bacterial Proteins; D004271:DNA, Fungal; C030069:spore-specific proteins, Bacillus