Figure 4 Phenotypes

of exponentially growing double mutant B. subtilis cells. A) dynA/floT double selleck mutant cells (note that membrane staining is highly heterogeneous between cells), white triangles indicate membrane abnormalities, B) mreB mutant cells grown in high magnesium medium, C-D) dynA/mreB double mutant cells growing in high magnesium medium. White or grey bars 2 μm. Figure 5 Growth curve of wild type cells (diamonds) or of dynA/ floT double mutant cells (squares) growing in S750 minimal glucose medium containing 0.1% casamino acids at 37° C. Data are means from four independently growing cultures. Based on its ability to tubulate membranes in vitro[11, 13], DynA may facilitate membrane invagination through a mechanical bending of the membrane, while FloT may be important to HDAC inhibitor generate a local environment favoring membrane curvature and/or recruitment of cell division proteins. In agreement with its function in lipid raft formation, a functional FloT-YFP fusion formed many discrete foci at the cell membrane [34] (Figure 3F). FloT-YFP was previously shown to move along random paths within or adjacent to the membrane [34]. These findings imply that due to the random movement, FloT would also be Wnt inhibitor frequently present at mid cell, which indeed was shown to be the case by colocalization of FloT-YFP with membrane

stain FM4-64 [34].To obtain a better idea about the extent of colocalization of FloT with the septal membrane, we quantified the number of FloT-YFP foci between cells. Indeed, 26% of FloT-YFP foci colocalized with the septal/polar membrane (184 foci analysed), or in other words, 22% of the cells had FloT-YFP fluorescence at the septum (Figure 3F, green FloT-YFP foci, red membrane) (148 cells analysed from 3 independent

experiments), showing Phosphoglycerate kinase that FloT is present at sites of cell division in a large fraction of the cells; even more cells contained FloT-YFP foci close to the cell centre. To investigate if one protein affects the localization of the other, we localized DynA-YFP in delta floT (yuaG) mutant cells. The localization pattern was indistinguishable from that of wild type cells (Figure 3G). Conversely, the absence of DynA did not visibly alter the localization pattern and dynamics of FloT-YFP (Figure 3H), showing that the proteins do not affect each other’s localization within the cell membrane and that they are not functionally linked. Synthetic phenotype of a dynA mreB double mutant strain Because floT dynA double mutant cells had a highly disturbed cell shape, we investigated the effect of a dynA deletion in combination with an mreB deletion. MreB is essential for the maintenance of rod shape in many bacteria, and the depletion of MreB leads to the generation of round cells that eventually lyse [20, 35].