DivIVA assembles into a ring-like structure at the polar septum during sporulation.

<p>(A) Schematic representation of sporulation in <i>Bacillus subtilis</i>. Depicted is a progenitor rod-shaped cell (above), the onset of asymmetric septation (middle), and formation of the polar septum (below), where the mother cell (MC) and forespore (FS) are labeled; activation of σ^F exclusively in the forespore is also depicted. (B) Immunoblot analysis of wild type cells (strain PY79; for genotypes, see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004526#pgen.1004526.s011" target="_blank">Table S1</a>), induced to sporulate and harvested at the times indicated above, using antisera specific to DivIVA or σ^A (an abundant unrelated protein that served as a loading control). Fraction of cells that had entered sporulation, as determined by epifluorescence microscopy, is indicated below. (C) Localization of DivIVA-GFP in strain KR604 induced to sporulate for 1.5 h. First panel: membranes were visualized using the fluorescent dye FM4-64; second panel: fluorescence from GFP; third panel: chromosomes were visualized using DAPI; fourth panel: overlay of membrane and GFP; fifth panel: overlay of membrane, GFP, and DNA. Cell #1 is a pre-divisional cell; cell #2 has elaborated a polar septum; cell #3 has just initiated asymmetric division, as evidenced by an increase in membrane staining at the future site of septation. Arrows indicate the sites of asymmetric division. (D) Top: rotation of the region of cell #3 indicated in (C), second panel, around the x- and y-axes. Degrees rotated are indicated below each panel; arrowheads indicate DivIVA patches. Bottom: GFP and DAPI fluorescence from cell #3 (C) represented as a three-dimensional surface. Arrow indicates DivIVA ring at the polar septum; arrowheads indicate the same DivIVA patches at poles. (E) Top: Localization of DivIVA-CFP (shown in green) and FtsZ-YFP (shown in red) in strain PE177. First panel: differential interference contrast (DIC); second panel: fluorescence from YFP; third panel: fluorescence from CFP; fourth panel: overlay of YFP and CFP; fifth panel: overlay of YFP, CFP, and DIC. Bottom: YFP and CFP fluorescence from the cell shown above represented as a three-dimensional surface and rotated to view at different angles. Panels 1–3: overlay of YFP and CFP. Arrow indicates DivIVA-CFP ring or constricting FtsZ-YFP at the polar division site; arrowheads indicate DivIVA-CFP patches at the extreme poles of the cell. Scale bars: 2 µm.