Figure 4.
SidA does not prevent assembly of the cell division machinery. (A) Subcellular localization of FtsZ was examined in a strain expressing ftsZ-eyfp from the vanillate-inducible promoter Pvan and overexpressing M2-sidA from a xylose-inducible promoter on a high-copy plasmid. Cells were grown to mid-exponential phase in rich medium with glucose and then shifted to xylose. Vanillate was added 1.5 h before shifting to xylose. At the times indicated, samples were taken and cells were imaged by DIC and epifluorescence microscopy (top two rows). The white asterisk indicates a FtsZ ring that is no longer associated with a constriction site and so likely moved and reassembled. (Bottom two rows) Localization of FtsN was examined in a strain expressing an egfp-ftsN fusion at its native chromosomal locus and overexpressing M2-sidA from a high-copy plasmid. The percentage of cells with foci after 4.5 h in xylose is shown below the last panel. (B) Localization of FtsI was examined in a strain expressing gfp-ftsI at its native chromosomal locus and overexpressing M2-sidA from a xylose-inducible promoter carried on a high-copy plasmid. Cells were grown to mid-exponential phase in rich medium with glucose and then placed on agarose pads containing xylose to produce M2-SidA. Individual cells were followed by time-lapse microscopy, with phase and epifluorescence images captured at the time points indicated. (C) Cells expressing gfp-ftsI and xylose-inducible M2-sidA were grown in the presence of xylose for 4.5 h to inhibit cell division and induce cellular filamentation. Cells were then placed on an agarose pad containing glucose to repress M2-sidA expression. Individual cells with localized GFP-FtsI were then followed by time-lapse microscopy, with phase and epifluorescence images captured at the time points indicated. Bars, 2 μm.