Division behavior and shape changes in isogenic ftsZ, ftsQ, ftsA, pbpB, and ftsE cell division mutants of Escherichia coli during temperature shift experiments

Abstract

Isogenic ftsZ, ftsQ, ftsA, pbpB, and ftsE cell division mutants of Escherichia coli were compared with their parent strain in temperature shift experiments. To improve detection of phenotypic differences in division behavior and cell shape, the strains were grown in glucose-minimal medium with a decreased osmolality (about 100 mosM). Already at the permissive temperature, all mutants, particularly the pbpB and ftsQ mutants,showed an increased average cell length and cell mass. The pbpB and ftsQ mutants also exhibited a prolonged duration of the constriction period. All strains, except ftsZ, continued to initiate new constrictions at 42rC, suggesting the involvement of FtsZ in an early step of the constriction process. The new constrictions were blunt in ftsQ and more pronounced in ftsA and pbpB filaments, which also had elongated median constrictions. Whereas the latter strains showed a slow recovery of cell division after a shift back to the permissive temperature, ftsZ and ftsQ filaments recovered quickly. Recovery of filaments occurred in all strains by the separation of newborn cells with an average length of two times L0, the length of newborn cells at the permissive temperature. The increased size of the newborn cells could indicate that the cell division machinery recovers too slowly to create normal-sized cells. Our results indicate a phenotypic resemblance between ftsA and pbpB mutants and suggest that the cell division gene products function in the order FtsZ-FtsQ-FtsA, PBP3. TheftsE mutant continued to constrict and divide at 42°C, forming short filaments, which recovered quickly after a shift back to the permissive temperature. After prolonged growth at 42°C, chains of cells, which eventually swelled up, were formed. Although theftsE mutant produced filaments in broth medium at the restrictive temperature, it cannot be considered a cell division mutant under the presently applied conditions. Escherichia coli is one of the very few organisms in which the genetics of the regulation of cell division can be studied: a number of specific conditional mutants that are affected in cell division have been described (for a review, see reference 11), and an enzymatic activity of one of the gene products has been demonstrated (18). Cell division in E. coli occurs by a seemingly simple ingrowth of the envelope layers without the help of additional structures like external wall bands (15) or internal skeletal compounds (30). In spite of this simplicity, it has appeared difficult to determine the specific role of the various cell division genes. First, the composition of the envelope layers at the division site has not yet been shown to differ chemically from the rest of the envelope. As a result, we have to describe the process in morphological terms and distinguish visual stages like initiation of cell wall ingrowth, formation of polar caps, and cell separation. The second reason is that inhibition of cell division can be the result of even slight perturbations of DNA synthesis (the SOS response), protein synthes...

Defining the rate-limiting processes of bacterial cytokinesis

“…Supporting this limiting role, we observed a significantly slowed septum closure rate in the presence of diminished septal transpeptidase activity by FtsI (MCI23 strain). We note that early studies have reported prolonged constriction periods caused by mutations in FtsI or FtsQ (114)(115)(116) or by overexpression of FtsN (116), although the corresponding cell wall synthesis activity was not assessed; these proteins are all essential components of the divisome involved in septal cell wall synthesis (117). Interestingly, recent work has shown that cytokinesis in fission yeast is primarily driven by cell wall synthesis rather than contraction of the force-generating actomyosin ring (118), suggesting that some similarities may exist in the mechanisms adapted by different kingdoms of life to coordinate cytokinesis in walled cells.…”

A model of membrane contraction predicting initiation and completion of bacterial cell division

“…In addition, in Escherichia coli, FtsZ-deficient mutants show no indentation of the membrane, whereas mutation of other essential division genes results in some level of indentation even when full division fails. 12 In liposomes seeded with membrane-tethered FtsZ, Z-ring structures are apparent below regions of indented membrane. 13 These results suggest that the force of indentation is generated by FtsZ, at least initially.…”

“…For a half life of recovery of 30 seconds, the corresponding observed rate of release from the membrane is 0.023 s −1 . Since the steady state of Z-ring assembly has been reached prior to the photobleaching, it follows that k bind2 p 1 (ī) (see Equations (12) and (16)) equals 0.023 s −1 (whereī is the average membrane-bound polymer length at equilibrium). With the dissociation constant κ fixed at 0.2 µM, it follows that the lower bounds of k bind1 and k bind2 are 0.142 µM −1 s −1 and 0.0284 s −1 , respectively.…”

“…Using fluorescence recovery after photobleaching (FRAP) the half life of scence in the Z-ring for wild-type FtsZ in E. coli was found to be 30 sec [29], corresponding te of release from the membrane of 0.023 s −1 . Since the steady state of Z-ring assembly has or to the photobleach, the observed release rate is given by k bind2 multiplied by the value uations (12) and (16)) whereī is the average bound polymer length at equilibrium. With onstant κ fixed at 0.2 µM, the model solution corresponding to an observed wild-type half 13 Fig.…”

A requirement for cell elongation protein RodZ and cell division proteins FtsN and DedD to maintain the small rod morphology of <i>Escherichia coli</i> at growth temperatures near 8°C

“…The other known reported E. coli mutant to form swollen prolate ellipsoids is the rodA ftsZ temperature-sensitive (Ts) double mutant at 42∞C (Begg and Donachie, 1985). The Ts mutation in rodA resulted in the formation of cocci due to a block in cell elongation, while the ftsZ mutation caused filaments due to a block in septal ring assembly (Begg and Donachie, 1985;Taschner et al, 1988). This swollen morphology is due to new peptidoglycan synthesis in the absence of cell elongation and septal ring formation (Begg and Donachie, 1985).…”