Envelope-associated nucleoid from Caulobacter crescentus stalked and swarmer cells

115

116

SummaryThe cell division protein FtsZ is composed of three regions based on sequence similarity: a highly conserved N-terminal region of Ϸ320 amino acids; a variable spacer region; and a conserved C-terminal region of eight amino acids. We show that FtsZ mutants missing different C-terminal fragments have dominant lethal effects because they block cell division in Caulobacter crescentus by two different mechanisms. Removal of the C-terminal conserved region, the linker, and 40 amino acids from the end of the Nterminal conserved region (FtsZ⌬C281) prevents the localization or the polymerization of FtsZ. Because two-hybrid analysis indicates that FtsZ⌬C281 does not interact with FtsZ, we hypothesize that FtsZ⌬C281 blocks cell division by competing with a factor required for FtsZ localization or that it titrates a factor required for the stability of the FtsZ ring. The removal of 24 amino acids from the C-terminus of FtsZ (FtsZ⌬C485) causes a punctate pattern of FtsZ localization and affects its interaction with FtsA. This suggests that the conserved C-terminal region of FtsZ is required for proper polymerization of FtsZ in Caulobacter and for its interaction with FtsA.

Section: Methodsmentioning

confidence: 99%

Section: Dna Manipulations Genetic Techniques Immunoblotting and Cementioning

confidence: 99%

Dominant C‐terminal deletions of FtsZ that affect its ability to localize in Caulobacter and its interaction with FtsA

Din

Quardokus

Sackett

et al. 1998

115

116

“…Caulobacter crescentus strains were grown at 30∞C in either in PYE complex-medium (Poindexter, 1964), or M2 minimal salts medium (Ely, 1991) supplemented with 0.2% glucose (M2G) or 0.1% glucose and 0.1% xylose (M2GX). The C. crescentus strains used were NA1000, a holdfast mutant derivative of wild-type strain CB15 (Evinger and Agabian, 1977); UJ945, a ftsH null mutant (Fischer et al, 2002); SG300, containing the groESL genes under the control of the P xylX promoter (see below) and SG400, containing the dnaKJ genes under the control of the P xylX promoter (see below). Plasmids used in this study are listed in Table 3.…”

Section: Bacterial Strains Growth Conditions and Plasmidsmentioning

confidence: 99%

Downregulation of the heat shock response is independent of DnaK and σ³² levels in Caulobacter crescentus

Silva

Simao

Susin

et al. 2003

“…Genetic techniques were carried out as described previously (Brun and Shapiro, 1992). Swarmer cells were isolated from exponential cultures grown in M2-G (Johnson and Ely, 1977), M2-GX or PYE (Poindexter, 1964) medium and processed as described previously (Evinger and Agabian, 1977). M2-GX is M2-G containing 0.3% (w/v) xylose.…”

Section: Dna Manipulations Genetic Techniques and Cell Synchronizationmentioning

confidence: 99%

Cell cycle and positional constraints on FtsZ localization and the initiation of cell division in Caulobacter crescentus

Quardokus

Din

Brun

2001

105

SummarySwarmer cells of Caulobacter crescentus are devoid of the cell division initiation protein FtsZ and do not replicate DNA. FtsZ is synthesized during the differentiation of swarmer cells into replicating stalked cells. We show that FtsZ first localizes at the incipient stalked pole in differentiating swarmer cells. FtsZ subsequently localizes at the mid-cell early in the cell cycle. In an effort to understand whether Z-ring formation and cell constriction are driven solely by the cell cycle-regulated increase in FtsZ concentration, FtsZ was artificially expressed in swarmer cells at a level equivalent to that found in predivisional cells. Immunofluorescence microscopy showed that, in these swarmer cells, simply increasing FtsZ concentration was not sufficient for Z-ring formation; Z-ring formation took place only in stalked cells. Expression of FtsZ in swarmer cells did not alter the timing of cell constriction initiation during the cell cycle but, instead, caused additional constrictions and a delay in cell separation. These additional constrictions were confined to sites close to the original mid-cell constriction. These results suggest that the timing and placement of Z-rings is tightly coupled to an early cell cycle event and that cell constriction is not solely dependent on a threshold level of FtsZ.