Bacillus subtilis is a Gram-positive microorganism which is able to differentiate during process called sporulation. A hallmark of sporulation in B. subtilis is the polar cell division. As occurs during vegetative cell division, the tubulin-like FtsZ forms a ring-like structure at mid-cell. At the onset of sporulation, however, the Z-ring migrates from mid-cell on a spiral trajectory to the two cell poles in a process that depends on the presence of SpoIIE. This protein colocalizes with the polar Z-rings. Asymmetric cell division otherwise appears to involve the same set of proteins as constitute the divisome during vegetative cell division. However, the resulting sporulation septum is much thinner. Accompanying these morphological changes is a coordinated programme of differential gene expression, involving intercellular signalling processes, that leads to the activation of the RNA polymerase sigma factors, sF and sG in the forespore and sE and sK in the mother cell [1].
Although, SpoIIE has a critical function in determining the site of formation of the sporulation septum, it is not understood (i) how it localises to the polar septum (ii) how it causes FtsZ to relocalise from mid-cell to the polar site (iii) what role SpoIIE plays in septal thinning, (iv) how its SpoIIAA~P phosphatase activity is controlled so that sF acitivation is delayed until the septum is completed (v) what role SpoIIE playes in SpoIIQ-SpoIIIAH channel formation.
SpoIIE from B. subtilis is an 827 residue protein that consists of three regions. It has 10 putative membrane-spanning segments (region I) at its amino terminus and a PP2C-type phosphatase domain (region III) at its C-terminus. The central region II is required for localisation of SpoIIE to the divisome and its reported interaction with FtsZ. The structure of the PP2C phosphatase domain of SpoIIE as well as a part of central domain together with phosphatase domain were already solved [2,3]. In contrast, the structure of N-terminal more than one thirds of SpoIIE and the character of its interactions with partner proteins are unknown. We recently identified a new partner of SpoIIE, the cytoskeletal protein, RodZ, which is essential for cell shape determination. This interaction is additionally required for asymmetric septum formation and sporulation.
We evaluated the positioning of the asymmetric septum and its accuracy by statistical analysis of the site of septation. We also clarified the role of SpoIIE, RefZ and MinCD on the accuracy of this process. In addition, we have employed a new method of "slimfield" microscopy to study the copy number, the oligomeric state and diffusion coefficient of SpoIIE in live cells and during its different roles in asymmetric septum site recognition, activation of sF and forespore engulfment.
This work was supported by Grant 2/0009/13 from the Slovak Academy of Sciences and by a Grant from the Slovak Research and Development Agency under contract APVV-14-0181.