Limits...
MukB colocalizes with the oriC region and is required for organization of the two Escherichia coli chromosome arms into separate cell halves.

Danilova O, Reyes-Lamothe R, Pinskaya M, Sherratt D, Possoz C - Mol. Microbiol. (2007)

Bottom Line: We show that in mukB mutant cells, the two chromosome arms do not separate into distinct cell halves, but extend from pole to pole with the oriC region located at the old pole.Mutations in topA, encoding topoisomerase I, do not suppress the aberrant positioning of chromosomal loci in mukB cells, despite suppressing the temperature-sensitivity and production of anucleate cells.We propose that MukBEF initiates the normal bidirectional organization of the chromosome from the oriC region.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, University of Oxford, Oxford OX1 3QU, UK.

ABSTRACT
The circular Escherichia coli chromosome is organized by bidirectional replication into two equal left and right arms (replichores). Each arm occupies a separate cell half, with the origin of replication (oriC) at mid-cell. E. coli MukBEF belongs to the ubiquitous family of SMC protein complexes that play key roles in chromosome organization and processing. In mukBEF mutants, viability is restricted to low temperature with production of anucleate cells, reflecting chromosome segregation defects. We show that in mukB mutant cells, the two chromosome arms do not separate into distinct cell halves, but extend from pole to pole with the oriC region located at the old pole. Mutations in topA, encoding topoisomerase I, do not suppress the aberrant positioning of chromosomal loci in mukB cells, despite suppressing the temperature-sensitivity and production of anucleate cells. Furthermore, we show that MukB and the oriC region generally colocalize throughout the cell cycle, even when oriC localization is aberrant. We propose that MukBEF initiates the normal bidirectional organization of the chromosome from the oriC region.

Show MeSH

Related in: MedlinePlus

Perturbation of <L-R> nucleoid organization in mukB cells. A. L3 and R3 positions by two-colour FISH. Analysis was restricted to four-foci cells having a normal nucleoid (as judged by DAPI staining). Foci from 123 MukB+ cells (AB1157) and 150 mukB cells (OS53) were binned into five positions from pole left (PL) to pole right (PR). The predominant four-focus patterns and the L3–R3 profiles are shown.B. Time-lapse analysis of R3 by FROS. Images of mukB cells (OS55) were taken every hour, kept at room temperature (G ∼5.5 h, as judged by cell elongation). Below, schematics show the predominant segregation pathways of R3 loci in wild-type and mukB cells. C. Same as B for R2 locus (OS30).
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2169520&req=5

fig02: Perturbation of <L-R> nucleoid organization in mukB cells. A. L3 and R3 positions by two-colour FISH. Analysis was restricted to four-foci cells having a normal nucleoid (as judged by DAPI staining). Foci from 123 MukB+ cells (AB1157) and 150 mukB cells (OS53) were binned into five positions from pole left (PL) to pole right (PR). The predominant four-focus patterns and the L3–R3 profiles are shown.B. Time-lapse analysis of R3 by FROS. Images of mukB cells (OS55) were taken every hour, kept at room temperature (G ∼5.5 h, as judged by cell elongation). Below, schematics show the predominant segregation pathways of R3 loci in wild-type and mukB cells. C. Same as B for R2 locus (OS30).

Mentions: The above results show that the <L-ori-R> organization about the transverse axis of normal cells is altered when MukBEF is absent, with polar ori positioning being reminiscent of the type of organization in Caulobacter crescentus (Viollier et al., 2004) and in chromosome 1 of Vibrio cholera (Fogel and Waldor, 2005). To further probe nucleoid organization, mukB cells grown at 22°C were examined using two-colour fluorescent in situ hybridization (FISH) with probes to L3 and R3 chromosome loci located on opposite arms at positions 2269 and 872 kb respectively close to the ter region (Fig. 2A). The patterns of L3–R3 positioning (Fig. 2A, top), and the L3 and R3 population profiles (bottom), were analysed in cells containing duplicated foci for each locus. These four-foci cells were enriched for by the addition of the cell division inhibitor cephalexin for 2.5 h (one-third of a generation time) prior to analysis (Wang et al., 2006). A <L-R> pattern in each sister nucleoid, arranged as <L3–R3–L3–R3> in pairs of sister nucleoids, was predominant (73%) for the wild-type strain, as reported earlier (Wang et al., 2006). This wild-type organization is also revealed by the L3–R3 population profile, in which L3 peaks at the left pole (PL) and at mid-cell, while R3 peaks at mid-cell and at the right pole (PR).


MukB colocalizes with the oriC region and is required for organization of the two Escherichia coli chromosome arms into separate cell halves.

Danilova O, Reyes-Lamothe R, Pinskaya M, Sherratt D, Possoz C - Mol. Microbiol. (2007)

Perturbation of <L-R> nucleoid organization in mukB cells. A. L3 and R3 positions by two-colour FISH. Analysis was restricted to four-foci cells having a normal nucleoid (as judged by DAPI staining). Foci from 123 MukB+ cells (AB1157) and 150 mukB cells (OS53) were binned into five positions from pole left (PL) to pole right (PR). The predominant four-focus patterns and the L3–R3 profiles are shown.B. Time-lapse analysis of R3 by FROS. Images of mukB cells (OS55) were taken every hour, kept at room temperature (G ∼5.5 h, as judged by cell elongation). Below, schematics show the predominant segregation pathways of R3 loci in wild-type and mukB cells. C. Same as B for R2 locus (OS30).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2169520&req=5

fig02: Perturbation of <L-R> nucleoid organization in mukB cells. A. L3 and R3 positions by two-colour FISH. Analysis was restricted to four-foci cells having a normal nucleoid (as judged by DAPI staining). Foci from 123 MukB+ cells (AB1157) and 150 mukB cells (OS53) were binned into five positions from pole left (PL) to pole right (PR). The predominant four-focus patterns and the L3–R3 profiles are shown.B. Time-lapse analysis of R3 by FROS. Images of mukB cells (OS55) were taken every hour, kept at room temperature (G ∼5.5 h, as judged by cell elongation). Below, schematics show the predominant segregation pathways of R3 loci in wild-type and mukB cells. C. Same as B for R2 locus (OS30).
Mentions: The above results show that the <L-ori-R> organization about the transverse axis of normal cells is altered when MukBEF is absent, with polar ori positioning being reminiscent of the type of organization in Caulobacter crescentus (Viollier et al., 2004) and in chromosome 1 of Vibrio cholera (Fogel and Waldor, 2005). To further probe nucleoid organization, mukB cells grown at 22°C were examined using two-colour fluorescent in situ hybridization (FISH) with probes to L3 and R3 chromosome loci located on opposite arms at positions 2269 and 872 kb respectively close to the ter region (Fig. 2A). The patterns of L3–R3 positioning (Fig. 2A, top), and the L3 and R3 population profiles (bottom), were analysed in cells containing duplicated foci for each locus. These four-foci cells were enriched for by the addition of the cell division inhibitor cephalexin for 2.5 h (one-third of a generation time) prior to analysis (Wang et al., 2006). A <L-R> pattern in each sister nucleoid, arranged as <L3–R3–L3–R3> in pairs of sister nucleoids, was predominant (73%) for the wild-type strain, as reported earlier (Wang et al., 2006). This wild-type organization is also revealed by the L3–R3 population profile, in which L3 peaks at the left pole (PL) and at mid-cell, while R3 peaks at mid-cell and at the right pole (PR).

Bottom Line: We show that in mukB mutant cells, the two chromosome arms do not separate into distinct cell halves, but extend from pole to pole with the oriC region located at the old pole.Mutations in topA, encoding topoisomerase I, do not suppress the aberrant positioning of chromosomal loci in mukB cells, despite suppressing the temperature-sensitivity and production of anucleate cells.We propose that MukBEF initiates the normal bidirectional organization of the chromosome from the oriC region.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, University of Oxford, Oxford OX1 3QU, UK.

ABSTRACT
The circular Escherichia coli chromosome is organized by bidirectional replication into two equal left and right arms (replichores). Each arm occupies a separate cell half, with the origin of replication (oriC) at mid-cell. E. coli MukBEF belongs to the ubiquitous family of SMC protein complexes that play key roles in chromosome organization and processing. In mukBEF mutants, viability is restricted to low temperature with production of anucleate cells, reflecting chromosome segregation defects. We show that in mukB mutant cells, the two chromosome arms do not separate into distinct cell halves, but extend from pole to pole with the oriC region located at the old pole. Mutations in topA, encoding topoisomerase I, do not suppress the aberrant positioning of chromosomal loci in mukB cells, despite suppressing the temperature-sensitivity and production of anucleate cells. Furthermore, we show that MukB and the oriC region generally colocalize throughout the cell cycle, even when oriC localization is aberrant. We propose that MukBEF initiates the normal bidirectional organization of the chromosome from the oriC region.

Show MeSH
Related in: MedlinePlus