The Caulobacter crescentus smc gene is required for cell cycle progression and chromosome segregation

Abstract

The highly conserved SMC (Structural Maintenance of Chromosomes) proteins function in chromosome condensation, segregation, and other aspects of chromosome dynamics in both eukaryotes and prokaryotes. A null mutation in the Caulobacter crescentus smc gene is conditionally lethal and causes a cell cycle arrest at the predivisional cell stage. Chromosome segregation in wild-type and smc null mutant cells was examined by monitoring the intracellular localization of the replication origin and terminus by using f luorescence in situ hybridization. In wild-type cells, the origin is located at the f lagellated pole of swarmer cells and, immediately after the initiation of DNA replication in stalked cells, one of the origins moves to the opposite pole, giving a bipolar localization of the origins. The terminus moves from the end of the swarmer cell opposite the origin to midcell. A subpopulation of the smc null mutant cells had mislocalized origins or termini, showing that the smc null mutation gives DNA segregation defects. Nucleoid morphology was also abnormal. Thus, we propose that the Caulobacter chromosomal origins have specific cellular addresses and that the SMC protein plays important roles in maintaining chromosome structure and in partitioning. The specific cell cycle arrest in the smc null mutant indicates the presence of a cell cycle checkpoint that senses perturbations in chromosome organization or segregation.

The bacterial nucleoid: A highly organized and dynamic structure

“…Both SMC and MukB were shown to cooperate with two accessory proteins, which regulate their function [Hirano and Hirano, 2004] and are essential for their activity in vivo [Yamanaka et al, 1996;Mascarenhas et al, 2002]. Inactivation of either of these complexes leads to temperaturesensitive growth due to a severe disturbance of nucleoid organization and chromosome segregation at elevated temperatures [Niki et al, 1991;Britton et al, 1998;Moriya et al, 1998;Jensen and Shapiro, 1999]. This phenotype was shown to be suppressed by mutations that cause excessive negative supercoiling, which suggests a role for these proteins in DNA compaction or folding [Sawitzke and Austin, 2000;Lindow et al, 2002a].…”

“…These problems are circumvented by a novel technique, which uses the lac or tet repressor fused to a fluorescent protein to specifically decorate tandem repeats of the cognate repressor binding site inserted at a chromosomal locus of interest Straight et al, 1996;Michaelis et al, 1997;Lau et al, 2003]. Using these two approaches, it was shown that bacteria possess mechanisms to orient the replication origin and terminus towards opposite poles in newborn cells Webb et al, 1997;Niki and Hiraga, 1998;Jensen and Shapiro, 1999;Fogel and Waldor, 2005]. This finding already indicates some degree of order within the nucleoid.…”

“…At the swarmer cell stage, which is characterized by a quiescent chromosome and, therefore, equivalent to the G 1 -phase of the cell cycle, the origin and terminus are located at the two opposite poles of the cell. After assembly of the replisome at the polar origin and initiation of DNA synthesis in the newly differentiated stalked cell, the duplicated strands are rapidly segregated, with one origin staying at the original position, while the second moves to the other end of the cell [Jensen and Shapiro, 1999;Jensen et al, 2001;Viollier et al, 2004]. As replication continues, the replisome is gradually displaced from its Viollier et al [2004] showed that the subcellular position of chromosomal sites is linearly dependent on their distance on the chromosomal map from the origin of replication.…”

Towards understanding the molecular basis of bacterial DNA segregation

“…Initiation of DNA replication results in the rapid formation of a second ParB focus at the opposite pole of the cell. This dynamic localization pattern coincides with the movement of the newly duplicated origin during the cell cycle (Jensen & Shapiro 1999). ParA localizes to the cell poles in C. crescentus but has not been assayed for polar oscillation in living cells (Mohl & Gober 1997).…”

Escherichia coli condensin MukB stimulates topoisomerase IV activity by a direct physical interaction

“…In addition to its structural similarity to canonical SMC proteins, MukB shares a common function with prokaryotic SMCs. Both E. coli mukB − strains and smc − strains from Bacillus subtilis and Caulobacter crescentus show temperature-sensitive colony formation and an increase in the number of anucleate cells at the permissive temperature, suggesting a deficiency in chromosome segregation (5,7,(15)(16)(17)(18)(19). Moreover, a convincing array of experiments has demonstrated that MukB functions as a condensin in vitro and in vivo (20)(21)(22)(23)(24)(25)(26)(27)(28).…”