Role of the C terminus of FtsK in Escherichia coli chromosome segregation

Abstract

FtsK is essential for Escherichia coli cell division. We report that cells lacking the C terminus of FtsK are defective in chromosome segregation as well as septation, often exhibiting asymmetrically positioned nucleoids and large anucleate regions. Combining the corresponding truncated ftsK gene with a mukB null mutation resulted in a synthetic lethal phenotype. When the truncated ftsK was combined with a minCDE deletion, chains of minicells were generated, many of which contained DNA. These results suggest that the C terminus of FtsK has an important role in chromosome partitioning.

FIG. 1

Localization of nucleoids and FtsZ in the ftsK1::cat mutant. Strain WM974 was grown in M9 medium supplemented with glucose and Casamino Acids (A to C and G to L) or Luria-Bertani medium (E and F) at 37°C. The wild-type parental control strain MG1655 was grown in Luria-Bertani medium at 37°C (D). Cells were grown to an optical density at 600 nm of approximately 0.2, fixed, stained with 0.5 μg of DAPI per ml, and visualized as described previously (13). Immunostaining with affinity-purified anti-FtsZ was also described previously (13). For panels D to F, cephalexin at 20 μg per ml was added to the culture, which was grown for an additional 2 h prior to fixation and DAPI staining. Panels A, D, E, F, G, and J show overlays of phase contrast plus DAPI staining (pseudocolored red); panels B, H, and K show overlays of phase contrast plus FtsZ immunostaining (green); and panels C, I, and L show overlays of DAPI (red) plus FtsZ immunostaining (green). Long arrows in panels A and B point to chains of cells with segregation defects, while short arrows in these panels highlight single cells with defects. Arrows in panels E and F point to filaments with severe segregation defects. Arrows in panel J highlight two cells, probably daughters, with misplaced nucleoids; the short arrow in panel K points to the FtsZ ring in the bottom cell that is off center relative to the cell and adjacent to the nucleoid; and the long arrows in panels K and L point to the FtsZ ring in the top cell that is asymmetric relative to the misplaced nucleoid but nevertheless at the cell midpoint. Bar, 5 μm.

FIG. 2

Chromosomal DNA in minicells of the ftsK1::cat strain. Strain WM975 (minB::kan ftsK1::cat) was grown in Luria-Bertani medium to an optical density at 600 nm of approximately 0.2. The cells were then fixed and stained with DAPI as described in the legend to Fig. 1. Panels A to C and D to F represent two different fields of cells, shown by phase contrast (A and D), DAPI staining (B and E), which appears bright on a dark background, and combined phase contrast and DAPI staining (C and F). Single arrows point to minicells that contain chromosomal DNA, while double arrows point to minicells lacking DNA. Bar, 5 μm.