RESUMEN

A discussion of some aspects of the regulation of chromosome replication, segregation and cell division in Escherichia coli.

Asunto(s)

Segregación Cromosómica , Proteínas del Citoesqueleto , Replicación del ADN , Proteínas de Escherichia coli , Escherichia coli/citología , Escherichia coli/genética , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Ciclo Celular/genética , División Celular/genética , Cromosomas Bacterianos , Regulación Bacteriana de la Expresión Génica

RESUMEN

Cell division normally follows the completion of each round of chromosome replication in Escherichia coli. Transcription of the essential cell division genes clustered at the mra region is shown here to depend on continuing chromosomal DNA replication. After chromosome replication was blocked by either nalidixic acid treatment or thymine starvation, the transcription of these cell division genes was repressed significantly. This suggests a way in which cell division is controlled by chromosome replication.

Asunto(s)

Cromosomas Bacterianos , Proteínas del Citoesqueleto , Replicación del ADN , Proteínas de Escherichia coli , Escherichia coli/citología , Escherichia coli/genética , Transcripción Genética , Proteínas Bacterianas/efectos de los fármacos , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , División Celular/efectos de los fármacos , División Celular/genética , Citoesqueleto/efectos de los fármacos , Replicación del ADN/efectos de los fármacos , Escherichia coli/efectos de los fármacos , Familia de Multigenes , Mutación , Ácido Nalidíxico/farmacología , Regiones Promotoras Genéticas , Rec A Recombinasas/efectos de los fármacos , Rec A Recombinasas/genética , Rec A Recombinasas/metabolismo , Proteínas Ribosómicas/efectos de los fármacos , Proteínas Ribosómicas/genética , Timidina/metabolismo

RESUMEN

The pattern of peptidoglycan (murein) segregation in cells of Escherichia coli with impaired activity of the morphogenetic proteins penicillin-binding protein 2 and RodA has been investigated by the D-cysteine-biotin immunolabeling technique (M. A. de Pedro, J. C. Quintela, J.-V. Höltje, and H. Schwarz, J. Bacteriol. 179:2823-2834, 1997). Inactivation of these proteins either by amdinocillin treatment or by mutations in the corresponding genes, pbpA and rodA, respectively, leads to the generation of round, osmotically stable cells. In normal rod-shaped cells, new murein precursors are incorporated all over the lateral wall in a diffuse manner, being mixed up homogeneously with preexisting material, except during septation, when strictly localized murein synthesis occurs. In contrast, in rounded cells, incorporation of new precursors is apparently a zonal process, localized at positions at which division had previously taken place. Consequently, there is no mixing of new and old murein. Old murein is preserved for long periods of time in large, well-defined areas. We propose that the observed patterns are the result of a failure to switch off septal murein synthesis at the end of septation events. Furthermore, the segregation results confirm that round cells of rodA mutants do divide in alternate, perpendicular planes as previously proposed (K. J. Begg and W. D. Donachie, J. Bacteriol. 180:2564-2567, 1998).

Asunto(s)

Proteínas Bacterianas/metabolismo , Proteínas Portadoras , Proteínas de Escherichia coli , Hexosiltransferasas/metabolismo , Proteínas de la Membrana , Complejos Multienzimáticos/metabolismo , Muramoilpentapéptido Carboxipeptidasa , Peptidoglicano/biosíntesis , Peptidil Transferasas/metabolismo , Amdinocilina/farmacología , Proteínas Bacterianas/genética , Escherichia coli/efectos de los fármacos , Escherichia coli/metabolismo , Escherichia coli/ultraestructura , Hexosiltransferasas/genética , Morfogénesis , Complejos Multienzimáticos/genética , Proteínas de Unión a las Penicilinas , Penicilinas/farmacología , Peptidil Transferasas/genética

RESUMEN

Resolution of chromosome dimers, by site-specific recombination between dif sites, is carried out in Escherichia coli by XerCD recombinase in association with the FtsK protein. We show here that a variety of altered FtsK polypeptides, consisting of the N-terminal (cell division) domain alone or with deletions in the proline-glutamine-rich part of the protein, or polypeptides consisting of the C-terminal domain alone are all unable to carry out dif recombination. Alteration of the putative nucleotide-binding site also abolishes the ability of FtsK to carry out recombination between dif sites.

Asunto(s)

Cromosomas Bacterianos , Escherichia coli/genética , Proteínas de la Membrana/metabolismo , Recombinación Genética , Proteínas de Escherichia coli , Eliminación de Secuencia

RESUMEN

Comparison of genome sequences from Eubacteria and Eukaryotic organelles shows that the order of genes in gene clusters encoding certain highly conserved cell division proteins and ribosomal proteins is itself highly conserved. Experiments with a cluster of cell division and related genes of E. coli have shown that this gene order is not essential for function. Comparisons between genomes also show that no pair of genes are necessarily adjacent in all genomes. The reason for the extreme conservation of order is therefore unknown, although one possible explanation might be the lateral exchange of tightly-linked groups of genes coding for co-adapted sets of proteins.

Asunto(s)

Bacterias/citología , Pared Celular/genética , Secuencia Conservada/genética , Células Eucariotas/citología , Orgánulos/genética , Proteínas Bacterianas/genética , División Celular/genética , Genes Bacterianos , Familia de Multigenes/genética , Ribosomas/genética , Homología de Secuencia de Ácido Nucleico

RESUMEN

Chromosome dimers, formed by homologous recombination between sister chromosomes, normally require cell division to be resolved into monomers by site-specific recombination at the dif locus of Escherichia coli. We report here that it is not in fact cell division per se that is required for dimer resolution but the action of the cytoplasmic domain of FtsK, which is a bifunctional protein required both for cell division and for chromosome partition.

Asunto(s)

Cromosomas Bacterianos , Proteínas de la Membrana/metabolismo , Sitios de Unión , Citoplasma/metabolismo , Dimerización , Proteínas de Escherichia coli , Proteínas de la Membrana/genética , Recombinación Genética

RESUMEN

The synthesis of the murein precursor lipid I is performed by MraY. We have shown that mraY is an essential gene for cell growth. Cells depleted of MraY first swell and then lyse. The expression of mraY DNA in vitro produces a 40-kDa polypeptide detectable by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.

Asunto(s)

Proteínas Bacterianas/genética , Escherichia coli/genética , Genes Bacterianos , Transferasas , Proteínas Bacterianas/aislamiento & purificación , Secuencia de Bases , División Celular/genética , Cartilla de ADN/genética , ADN Bacteriano/genética , Escherichia coli/citología , Escherichia coli/crecimiento & desarrollo , Expresión Génica , Prueba de Complementación Genética , Mutagénesis Insercional , Fenotipo , Plásmidos/genética , Transferasas (Grupos de Otros Fosfatos Sustitutos)

Asunto(s)

Cromosomas Bacterianos , Proteínas de la Membrana/fisiología , División Celular , Escherichia coli/genética , Proteínas de Escherichia coli , Regulación Bacteriana de la Expresión Génica , Proteínas de la Membrana/genética , Regulón , Respuesta SOS en Genética

RESUMEN

Deletion of ftsK results in the inhibition of cell division, but this inhibition can be reversed by a plasmid carrying only the first approximately 17% of ftsK. The division block can be suppressed in most mutants by deletion of dacA, which codes for the D-alanine:D-alanine carboxypeptidase PBP5, or in all mutants by overexpression of ftsN. Overexpression of ftsK inhibits cell division and the formation of FtsZ rings. This division block is not due to the induction of either the SOS or the heat shock regulons.

Asunto(s)

Proteínas Bacterianas/fisiología , División Celular/fisiología , Proteínas de la Membrana/fisiología , Proteínas Bacterianas/química , Secuencia de Bases , Cartilla de ADN , Proteínas de Escherichia coli , Proteínas de la Membrana/química , Mutación , Plásmidos , Regiones Promotoras Genéticas , Respuesta SOS en Genética , Eliminación de Secuencia

RESUMEN

The localization of FtsI (PBP3), a penicillin-binding protein specifically required for cell division in Escherichia coli, was investigated by immunofluorescence microscopy and found to localize to the septum. The localization of FtsI was not observed in ftsZ or ftsA mutants, indicating that it was dependent on the prior localization of these proteins. Addition of furazlocillin, a specific inhibitor of FtsI, prevented localization of FtsI even though FtsZ and FtsA localization occurred. Interestingly, the localization of FtsN was also prevented by furazlocillin. FtsZ displayed limited localization in furazlocillin-treated cells, whereas it was efficiently localized in FtsI-depleted cells. FtsW, another essential cell division protein, was also localized to the septum.

Asunto(s)

Proteínas Bacterianas/análisis , Proteínas Portadoras , Proteínas del Citoesqueleto , Proteínas de Escherichia coli , Escherichia coli/química , Hexosiltransferasas/análisis , Imidazolidinas , Proteínas de la Membrana , Complejos Multienzimáticos/análisis , Muramoilpentapéptido Carboxipeptidasa , Peptidil Transferasas/análisis , Azlocilina/análogos & derivados , Azlocilina/farmacología , Proteínas Bacterianas/genética , División Celular/efectos de los fármacos , Escherichia coli/genética , Hexosiltransferasas/antagonistas & inhibidores , Complejos Multienzimáticos/antagonistas & inhibidores , Mutación , Proteínas de Unión a las Penicilinas , Penicilinas/farmacología , Peptidil Transferasas/antagonistas & inhibidores

RESUMEN

In the spherical cells of Escherichia coli rodA mutants, division is initiated at a single point, from which a furrow extends progressively around the cell. Using "giant" rodA ftsA cells, we confirmed that each new division furrow is initiated at the midpoint of the previous division plane and runs perpendicular to it.

Asunto(s)

Escherichia coli/citología , División Celular , Polaridad Celular , Morfogénesis

RESUMEN

Increasing FtsZ induces the formation of minicells at cell poles but does not increase the frequency or timing of central divisions. A coordinate increase in both FtsZ and FtsA, however, increases the frequency of both polar and central divisions.

Asunto(s)

Proteínas Bacterianas/metabolismo , Proteínas del Citoesqueleto , Proteínas de Escherichia coli , Escherichia coli/citología , Proteínas Bacterianas/genética , División Celular , Proteínas de la Membrana/genética , Proteínas Recombinantes/metabolismo

RESUMEN

In the absence of exogenous promoters, plasmid-mediated complementation of the temperature-sensitive ftsW201 allele requires the presence of the full coding sequence of ftsW plus upstream DNA encompassing the C-terminus of mraY and the full coding sequence of murD. We used molecular and genetic techniques to introduce an insertional inactivation into the chromosomal copy of ftsW, in the presence of the plasmid-borne wild-type ftsW gene under the control of P(BAD). In the absence of arabinose, the ftsW-null strain is not viable, and a shift from arabinose- to glucose-containing liquid medium resulted in a block in division, followed by cell lysis. Immunofluorescence microscopy revealed that in ftsW-null filaments, the FtsZ ring is absent in 50-60% of filaments, whilst between one and three Z-rings per filament can be detected in the remainder of the population, with the majority of these containing only one Z-ring per filament. We also demonstrated that the expression of only ftsWS (the smaller of two ftsW open reading frames) from P(BAD) is sufficient for complementation of the ftsW-null allele. We conclude that FtsW is an essential cell-division protein in Escherichia coli, and that it plays a role in the stabilization of the FtsZ ring during cell division.

Asunto(s)

Proteínas Bacterianas/genética , Proteínas del Citoesqueleto , Escherichia coli/genética , Proteínas de la Membrana , Alelos , Proteínas Bacterianas/metabolismo , División Celular/genética , Cromosomas Bacterianos , Escherichia coli/crecimiento & desarrollo , Escherichia coli/metabolismo , Eliminación de Gen , Expresión Génica , Genes Bacterianos , Prueba de Complementación Genética , Mutagénesis Insercional , Fenotipo , Regiones Promotoras Genéticas

RESUMEN

Two new mutations in the cell division gene ftsW have been isolated and characterized. The ftsW263(Ts) mutation results in a block to division at the initiation stage, similar to that previously observed with the ftsW201(Ts) mutation. The ftsW1640(Ts) mutation, however, causes a block to division at a later stage. The ftsW201 and ftsW263 mutants were shown to be phenotypically sensitive to the genetic background and growth conditions and are possibly relA dependent. Immunofluorescence microscopy showed that the FtsZ protein can localize to presumptive division sites in strains carrying ftsW(Ts) mutations at the nonpermissive temperature, suggesting that FtsW is unlikely to be specifically required for the localization of FtsZ to the division site. Examination of the localization of FtsZ in an ftsW rodA double mutant (lemon-shaped cells) revealed several classes of cells ranging from a common class where an FtsZ ring structure is absent to a class where FtsZ forms a complete ring at the midpoint of a lemon-shaped cell, suggesting a role for FtsW in the establishment of a stable FtsZ-based septal structure. We further demonstrate that two FtsW peptides, FtsWL (large) and FtsWS (small), can be identified and that the expression of ftsWS is sufficient for complementation of ftsW(Ts) mutations.

Asunto(s)

Proteínas Bacterianas/genética , Proteínas Bacterianas/aislamiento & purificación , Proteínas del Citoesqueleto , Proteínas de Escherichia coli , Escherichia coli/citología , Proteínas de Unión al GTP/genética , Proteínas de la Membrana , Sistemas de Lectura , Alelos , Secuencia de Aminoácidos , Proteínas Bacterianas/inmunología , Compartimento Celular , División Celular/genética , Escherichia coli/genética , Técnica del Anticuerpo Fluorescente , Ligasas/genética , Datos de Secuencia Molecular , Mutación , Fenotipo , Análisis de Secuencia de ADN

RESUMEN

The phenotype of a minC mutant has been reexamined and found to correspond closely to the quantitative predictions of Teather et al. (R. M. Teather, J. F. Collins, and W. D. Donachie, J. Bacteriol. 118:407-413, 1974). We confirm that the number of septa formed per generation per cell length is fixed and independent of the number of available division sites and that "division potential" is directly proportional to cell length. In the minC mutant, septa form with equal probabilities at cell poles, cell centers, and cell quarters. In addition, we show that the time to next division is inversely related to cell length while division is asynchronous in long cells, suggesting that a single cell can form only one septum at a time.

Asunto(s)

Proteínas Bacterianas/genética , Escherichia coli/citología , Escherichia coli/genética , Mutación , División Celular , Análisis de Regresión , Factores de Tiempo

RESUMEN

A mutation in a newly discovered Escherichia coli cell division gene, ftsK, causes a temperature-sensitive late-stage block in division but does not affect chromosome replication or segregation. This defect is specifically suppressed by deletion of dacA, coding for the peptidoglycan DD-carboxypeptidase, PBP 5. FtsK is a large polypeptide (147 kDa) consisting of an N-terminal domain with several predicted membrane-spanning regions, a proline-glutamine-rich domain, and a C-terminal domain with a nucleotide-binding consensus sequence. FtsK has extensive sequence identity with a family of proteins from a wide variety of prokaryotes and plasmids. The plasmid proteins are required for intercellular DNA transfer, and one of the bacterial proteins (the SpoIIIE protein of Bacillus subtilis) has also been implicated in intracellular chromosomal DNA transfer.

Asunto(s)

Proteínas Bacterianas , Escherichia coli/crecimiento & desarrollo , Escherichia coli/genética , Genes Bacterianos , Hexosiltransferasas , Proteínas de la Membrana/genética , Peptidil Transferasas , Secuencia de Aminoácidos , Secuencia de Bases , Sitios de Unión , Proteínas Portadoras/genética , División Celular/genética , Mapeo Cromosómico , Secuencia de Consenso , Escherichia coli/citología , Proteínas de Escherichia coli , Prueba de Complementación Genética , Datos de Secuencia Molecular , Muramoilpentapéptido Carboxipeptidasa/genética , Mutación , Proteínas de Unión a las Penicilinas , Peptidoglicano/biosíntesis , Fenotipo , Estructura Secundaria de Proteína , Recombinación Genética , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Supresión Genética

RESUMEN

In the rod-shaped cells of E. coli, chromosome segregation takes place immediately after replication has been completed. A septum then forms between the two sister chromosomes. In the absence of certain membrane proteins, cells grow instead as large, multichromosomal spheres that divide successively in planes that are at right angles to one another. Although multichromosomal, the spherical cells cannot be maintained as heterozygotes. These observations imply that, in these mutants, each individual chromosome gives rise to a separate clone of descendant cells. This suggests a model in which sites for cell division form between pairs of sister chromosomes at the time of segregation, but are not used in spherical cells until further rounds of replication have taken place, thus ensuring clonal ('hierarchical') segregation of chromosomes into progeny cells. The role of the morphogenetic membrane proteins is to convert the basically spherical cell into a cylinder that is able to divide as soon as replication and segregation have been completed, and thus to maximise the number of viable cells per genome.

Asunto(s)

Ciclo Celular , Cromosomas Bacterianos/fisiología , Cromosomas Bacterianos/ultraestructura , Proteínas de Escherichia coli , Escherichia coli/citología , Escherichia coli/genética , Proteínas de la Membrana , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , División Celular , Replicación del ADN , Fase G2 , Genes Bacterianos , Modelos Biológicos

RESUMEN

The product of the ftsW gene has been identified as a polypeptide that, like the related RodA protein, shows anomalous mobility on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. FtsW is produced at low levels that can be increased by altering the translation initiation region of the mRNA. Overproduction of FtsW strongly inhibits cell growth. A new mutant allele, ftsW201, causes a temperature-dependent block in the initiation stage of cell division which is similar to the division block in ftsZ mutants. The block in initiation of division in the ftsW201 allele is shown to be independent of FtsZ or the FtsZ inhibitor, SulA. In addition, the ftsW201 mutant is hypersensitive to overproduction of the division initiation protein FtsZ at the permissive temperature. Our results suggest a role for FtsW in an early stage of division which may involve an interaction with FtsZ.

Asunto(s)

Proteínas Bacterianas/genética , Proteínas del Citoesqueleto , Proteínas de Escherichia coli , Escherichia coli/crecimiento & desarrollo , Proteínas de la Membrana , Alelos , Proteínas Bacterianas/biosíntesis , Secuencia de Bases , División Celular/genética , Escherichia coli/citología , Escherichia coli/genética , Microscopía , Datos de Secuencia Molecular , Mutación , Fenotipo , Biosíntesis de Proteínas , Análisis de Secuencia de ADN , Supresión Genética

RESUMEN

A 490-bp DNA segment spanning the junction between the ftsA and ftsZ genes inhibits cell division when present in high copy number. We show that this segment contains an antisense promoter and an antisense transcription terminator which define a new gene, stfZ.

Asunto(s)

Proteínas Bacterianas/biosíntesis , Proteínas del Citoesqueleto , ADN sin Sentido , Proteínas de Escherichia coli , Escherichia coli/genética , Genes Bacterianos , Regiones Promotoras Genéticas , Transcripción Genética , Secuencia de Aminoácidos , Proteínas Bacterianas/genética , Secuencia de Bases , División Celular/genética , Escherichia coli/citología , Datos de Secuencia Molecular , Mapeo Restrictivo

RESUMEN

For normal cell division, the ftsZ gene must be transcribed from a number of promoters that are located within the proximal upstream genes (ddlB, ftsQ, and ftsA). We show that the main promoters have identical responses to changes in growth rate, i.e., under all conditions, the frequency of transcription per septum formed is approximately constant and independent of cell size or growth rate per se. We also show that transcription from these promoters is independent of stationary-phase transcription factor sigma s.

Asunto(s)

Proteínas Bacterianas/genética , Proteínas del Citoesqueleto , Escherichia coli/crecimiento & desarrollo , Proteínas de Unión al GTP/genética , Regulación Bacteriana de la Expresión Génica , Regiones Promotoras Genéticas/genética , Proteínas Bacterianas/biosíntesis , Proteínas Bacterianas/metabolismo , División Celular , Escherichia coli/genética , Proteínas de Unión al GTP/biosíntesis , Factor sigma/metabolismo , Transcripción Genética , beta-Galactosidasa/biosíntesis