Characterization of a prokaryotic SMC protein involved in chromosome partitioning

Abstract

smc of Bacillus subtilis encodes a homolog of eukaryotic SMC proteins involved in chromosome condensation, pairing, and partitioning. A null mutation in B. subtilis smc caused a temperature-sensitive-lethal phenotype in rich medium. Under permissive conditions, the mutant had abnormal nucleoids, ∼10% of the cells were anucleate, and assembly of foci of the chromosome partitioning protein Spo0J was altered. In combination with a null mutation in spo0J, the smc mutation caused a synthetic phenotype; cell growth was slower and ∼25% of the cells were anucleate. Our results demonstrate that the B. subtilis Smc protein, like its eukaryotic counterpart, plays an important role in chromosome structure and partitioning.

Penicillin-Binding Protein-Related Factor A Is Required for Proper Chromosome Segregation in Bacillus subtilis

“…However, LP101 cultures grown without xylose contained significantly more anucleate cells (ϳ24%) than did cultures grown in the presence of xylose (ϳ15% [Table 3]), suggesting that the lack of PrfA exacerbates the smc phenotype. Cultures of smc (LP99) cells grown under similar conditions contained ϳ13% anucleate cells (Table 3), which is consistent with previous reports (ϳ10 to 15% anucleate cells [6,26]). We also examined the LP101 cells grown to stationary phase in PAB medium with or without xylose.…”

“…The smc gene (for "structural maintenance of chromosomes") is required for chromosome structure and partitioning in B. subtilis and inactivation of smc produces cells with chromosome segregation defects similar to but more severe than those observed for prfA cells. For example, smc null mutant cells grown at 23 or 30°C contain about 10 to 15% anucleate cells compared to ϳ0.9 to 3% for prfA cells (Table 3) (6,26). We attempted to generate a prfA smc double mutant by transforming PS2061 (⌬prfA::spc) cells with chromosomal DNA from strain RB35 (⌬smc::kan) and selecting transformants on LB plates supplemented with kanamycin at 30°C (strain RB35 is temperature sensitive for growth in rich medium [6]).…”

“…For example, smc null mutant cells grown at 23 or 30°C contain about 10 to 15% anucleate cells compared to ϳ0.9 to 3% for prfA cells (Table 3) (6,26). We attempted to generate a prfA smc double mutant by transforming PS2061 (⌬prfA::spc) cells with chromosomal DNA from strain RB35 (⌬smc::kan) and selecting transformants on LB plates supplemented with kanamycin at 30°C (strain RB35 is temperature sensitive for growth in rich medium [6]). Although we obtained a number of very small colonies after 2 days of incubation, detailed analysis of several of these transformants suggested that they most probably contained suppressor mutations (results not shown).…”

Independent Segregation of the Two Arms of theEscherichia coli oriRegion Requires neither RNA Synthesis nor MreB Dynamics

“…Therefore, the key to efficient and faithful segregation is likely to reside in chromosome organization itself and the processes that drive this organization, as well as independent replication by spatially separated replisomes tracking along the DNA and the subsequent decatenation by TopoIV. Consistent with this view, aberrant chromosome organization as a consequence of absence of functional SMC complexes (MukbEF) leads to an altered pattern of replication-segregation and to failures in chromosome segregation (8,10,44). It seems to us that the independent tracking of sister replisomes along DNA, outward from oriC (47), may facilitate the segregation of newly replicated sister chromosomes into separate cell halves, thereby allowing the entropic mechanism to mediate the segregation process efficiently.…”

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].…”