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Category: Bacterial Pathogenesis
Chromosome Replication and Segregation, Page 1 of 2
< Previous page | Next page > /docserver/preview/fulltext/10.1128/9781555817992/9781555812058_Chap07-1.gif /docserver/preview/fulltext/10.1128/9781555817992/9781555812058_Chap07-2.gifAbstract:
This chapter reviews our current understanding of bacterial DNA replication and chromosome partitioning in Bacillus subtilis and makes comparisons to Escherichia coli and other organisms where appropriate. Bacterial chromosome replication initiates once per cell division cycle in response to a signal that is tightly coupled to cell mass. Although the helicase function of B. subtilis DnaC has not yet been confirmed biochemically, two types of dna(TS) mutations, defective in initiation and elongation, map to dnaC. Strong interaction between the helicase and the primase has been demonstrated in Bacillus stearotrtermophilus. The current understanding of bacterial chromosome partitioning can be simplified into three steps: (i) origin region separation and repositioning, (ii) overall chromosome organization and compaction, and (iii) terminus region separation. This final step includes chromosome decatenation, chromosome dimer-to-monomer resolution when necessary, and movement of the termini to either side of midcell before completion of medial division. The structural maintenance of chromosomes (SMC) protein family is well conserved and is important for chromosome segregation in bacteria, archaea, and eukaryotes. Both B. subtilis and E. coli have proteins that appear to be involved in postseptational chromosome partitioning. These proteins, SpoIIIE and FtsK, respectively, have domains that are homologous to the DNA translocation domains of proteins involved in conjugative plasmid transfer. In B. subtilis and Caulobacter crescentus, SMC functions in chromosome partitioning presumably by affecting chromosome organization and compaction. All organisms seem to have proteins that contribute to chromosome folding and compaction.
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Simplified cartoon of the bacterial cell cycle and chromosome orientation. The chromosome is indicated by a thin oval inside the cell. The origin of replication (oriC) is indicated by a small gray circle, and the terminus of replication (ter) is indicated by a gray square. The replisome is indicated by two triangles, one for each replication fork that initiates from oriC. In this model, replication initiates at or near midcell, the origins rapidly separate, replication continues as the newly replicated DNA is refolded (in part by Smc), and copies separate from each other. The cell division machinery assembles at midcell, and cells divide. This is simplified, because at rapid growth rates, newborn cells have a partly duplicated chromosome and the origin regions have already duplicated and separated to the cell quarters.
Simplified cartoon of the bacterial cell cycle and chromosome orientation. The chromosome is indicated by a thin oval inside the cell. The origin of replication (oriC) is indicated by a small gray circle, and the terminus of replication (ter) is indicated by a gray square. The replisome is indicated by two triangles, one for each replication fork that initiates from oriC. In this model, replication initiates at or near midcell, the origins rapidly separate, replication continues as the newly replicated DNA is refolded (in part by Smc), and copies separate from each other. The cell division machinery assembles at midcell, and cells divide. This is simplified, because at rapid growth rates, newborn cells have a partly duplicated chromosome and the origin regions have already duplicated and separated to the cell quarters.
Model of initiation mechanism of B. subtilis chromosome replication. The dnaA gene constitutes an operon with dnaN ( 101 ). DnaA boxes exist in two noncoding regions upstream and downstream of dnaA, where DnaA actually binds ( 29 ). The dnaA-dnaN operon is probably expressed after initiation of replication ( 100 ) and is autoregulated by DnaA ( 101 ). Thus, DnaA newly synthesized after initiation would stop its transcription. Two DnaA box clusters are required for initiation of replication in vivo (shown as ars, autonomously replicating sequence) ( 86 ), and the regions form a loop mediated by DnaA in vitro ( 57 ). It is unclear whether the loop is formed only at the time of initiation. This loop formation opens double-stranded DNA locally at an AT-rich sequence between dnaA and dnaN ( 57 ), consistent with in vivo observations that oriC plasmid and chromosome replication start at this non-coding region ( 87 , 90 ). DnaB, DnaD, and Dnal are probably components of a primosome ( 14 ) and thus play roles for loading the DnaC helicase into the unwound region. DnaD interacts with DnaA ( 45 ), but the role of the interaction is still unclear. DnaB exhibits single-stranded DNA-binding activity ( 128 ) and forms an oligomer ( 45 ), similar to E. coli DnaC helicase loader. However, DnaB did not interact with DnaA, DnaC, DnaD, or Dnal by the yeast two-hybrid assay ( 45 ). Its precise role is still obscure. Dnal interacted strongly with the DnaC helicase ( 42 ), indicating that it acts as a component of the helicase loading system. Once the helicase is loaded into oriC, primase (DnaG) and τ subunit (DnaX) of DNA polymerase III are assembled by protein-protein interaction followed by formation of the replisome on oriC.
Model of initiation mechanism of B. subtilis chromosome replication. The dnaA gene constitutes an operon with dnaN ( 101 ). DnaA boxes exist in two noncoding regions upstream and downstream of dnaA, where DnaA actually binds ( 29 ). The dnaA-dnaN operon is probably expressed after initiation of replication ( 100 ) and is autoregulated by DnaA ( 101 ). Thus, DnaA newly synthesized after initiation would stop its transcription. Two DnaA box clusters are required for initiation of replication in vivo (shown as ars, autonomously replicating sequence) ( 86 ), and the regions form a loop mediated by DnaA in vitro ( 57 ). It is unclear whether the loop is formed only at the time of initiation. This loop formation opens double-stranded DNA locally at an AT-rich sequence between dnaA and dnaN ( 57 ), consistent with in vivo observations that oriC plasmid and chromosome replication start at this non-coding region ( 87 , 90 ). DnaB, DnaD, and Dnal are probably components of a primosome ( 14 ) and thus play roles for loading the DnaC helicase into the unwound region. DnaD interacts with DnaA ( 45 ), but the role of the interaction is still unclear. DnaB exhibits single-stranded DNA-binding activity ( 128 ) and forms an oligomer ( 45 ), similar to E. coli DnaC helicase loader. However, DnaB did not interact with DnaA, DnaC, DnaD, or Dnal by the yeast two-hybrid assay ( 45 ). Its precise role is still obscure. Dnal interacted strongly with the DnaC helicase ( 42 ), indicating that it acts as a component of the helicase loading system. Once the helicase is loaded into oriC, primase (DnaG) and τ subunit (DnaX) of DNA polymerase III are assembled by protein-protein interaction followed by formation of the replisome on oriC.
Model of proteins present at the replication fork of B. subtilis.
Model of proteins present at the replication fork of B. subtilis.
Spo0J binding sites on the B. subtilis chromosome. oriC is at 0°/360° on the circular chromosome. The eight known par sites ( 70 ) are indicated.
Spo0J binding sites on the B. subtilis chromosome. oriC is at 0°/360° on the circular chromosome. The eight known par sites ( 70 ) are indicated.
Model of B. subtilis SMC. SMC is an antiparallel homodimer with two long coiled-coil regions separated by a flexible hinge ( 82 ).
Model of B. subtilis SMC. SMC is an antiparallel homodimer with two long coiled-coil regions separated by a flexible hinge ( 82 ).
Chromosome partitioning events specific to the terminus region. (A) Chromosome decatenation. (B) When necessary, a dif site-specific recombination resolves a chromosome dimer (left) to two monomers. (C) Model for SpoIIIE (or FtsK) movement of trapped chromosome out of the division septum.
Chromosome partitioning events specific to the terminus region. (A) Chromosome decatenation. (B) When necessary, a dif site-specific recombination resolves a chromosome dimer (left) to two monomers. (C) Model for SpoIIIE (or FtsK) movement of trapped chromosome out of the division septum.
B. subtilis genes involved in chromosomal DNA replication
a The numbers indicate nucleotide positions of each gene (coding region) on the whole genome. To the left and right of the hyphen are positions of the first and last letters of start and stop codons, respectively. (See http://bacillus.genome.ad.jp/ or http://genolist.pasteur.fr/SubtiList/)
b Personal communication from S.D. Ehrlich.
c dnaA and dnaN constitute an operon ( 101 ). dnal is a member of the dnaB operon ( 13 ). lig and pcrA belong to a putative operon ( 103 ).
d E. coli and Thermus thermophilus, γ subunit is also produced from the dnaX gene by translational frameshifting and transcriptional slippage, respectively ( 59 ). The existence of γ has not yet been confirmed in B. subtilis.
e The core of E. coli DNA polymerase III consists of α, є, and θ subunits ( 5 ). In B. subtilis, the activity of proofreading exonuclease (є) is included in the α subunit, and no homologs of θ are found.
B. subtilis genes involved in chromosomal DNA replication
a The numbers indicate nucleotide positions of each gene (coding region) on the whole genome. To the left and right of the hyphen are positions of the first and last letters of start and stop codons, respectively. (See http://bacillus.genome.ad.jp/ or http://genolist.pasteur.fr/SubtiList/)
b Personal communication from S.D. Ehrlich.
c dnaA and dnaN constitute an operon ( 101 ). dnal is a member of the dnaB operon ( 13 ). lig and pcrA belong to a putative operon ( 103 ).
d E. coli and Thermus thermophilus, γ subunit is also produced from the dnaX gene by translational frameshifting and transcriptional slippage, respectively ( 59 ). The existence of γ has not yet been confirmed in B. subtilis.
e The core of E. coli DNA polymerase III consists of α, є, and θ subunits ( 5 ). In B. subtilis, the activity of proofreading exonuclease (є) is included in the α subunit, and no homologs of θ are found.
Orthologous proteins involved in chromosome replication in gram-positive bacteria and E. coli a
a Orthologous genes were searched by BLAST 2.0 (Advanced) and specialized BLAST to Microbial Genomes (finished and unfinished) at the National Center for Biotechnology Information website using B. subtilis proteins as query. Where there were no orthologs in B. subtilis, E. coli proteins were used as query. The amino acid sequences of query proteins were obtained from http://bacillus.genome.ad.jp/ and http://dna.aist-nara.ac.jp/ecoli/ for B. subtilis and E. coli, respectively. Orthologous genes having alignment scores of >100 are listed except where noted. In Spy, Sau, Efa, and Cdi, the presence of the orthologs is shown as “+” because the gene names were not available in the databases. When orthologs were not found but the genome sequencing had not yet been completed, the columns remain blank. Genome sequencing has finished in Cac, but annotation of genes has not been done. Therefore, when orthologs are not detected, “−” is given in such columns.
b Abbreviations of strains, with references: Eco, E. coli; Bsu, B. subtilis; Mpn, Mycoplasma pneumoniae ( 36 ); Spy, Streptococcus pyogenes; Sau, Staphylococcus aureus ( 2 , 102 ); Mtu, Mycobacterium tuberculosis ( 19 , 114 ); Cac, Clostridium acetobutylicum ( 116 , 132 ); Efa, Enterococcus faecalis; Cdi, Corynebacterium diphtheriae; Sco, Streptomyces coelicolor ( 17 , 27 , 110 ); Dra, Deinococcus radiodurans ( 139 ).
c Alignment score, 30–50.
d BLAST search (tblastn) identified a homologous gene here, but no coding sequence is assigned in the database.
e Alignment score, 50–100.
f In this organism, δ has been recently found, and a τδδ′ complex (without γ) actually acts as a clamp loader ( 16 ).
g These genes are named dnaE in their original databases but are renamed dnaG in this table.
Orthologous proteins involved in chromosome replication in gram-positive bacteria and E. coli a
a Orthologous genes were searched by BLAST 2.0 (Advanced) and specialized BLAST to Microbial Genomes (finished and unfinished) at the National Center for Biotechnology Information website using B. subtilis proteins as query. Where there were no orthologs in B. subtilis, E. coli proteins were used as query. The amino acid sequences of query proteins were obtained from http://bacillus.genome.ad.jp/ and http://dna.aist-nara.ac.jp/ecoli/ for B. subtilis and E. coli, respectively. Orthologous genes having alignment scores of >100 are listed except where noted. In Spy, Sau, Efa, and Cdi, the presence of the orthologs is shown as “+” because the gene names were not available in the databases. When orthologs were not found but the genome sequencing had not yet been completed, the columns remain blank. Genome sequencing has finished in Cac, but annotation of genes has not been done. Therefore, when orthologs are not detected, “−” is given in such columns.
b Abbreviations of strains, with references: Eco, E. coli; Bsu, B. subtilis; Mpn, Mycoplasma pneumoniae ( 36 ); Spy, Streptococcus pyogenes; Sau, Staphylococcus aureus ( 2 , 102 ); Mtu, Mycobacterium tuberculosis ( 19 , 114 ); Cac, Clostridium acetobutylicum ( 116 , 132 ); Efa, Enterococcus faecalis; Cdi, Corynebacterium diphtheriae; Sco, Streptomyces coelicolor ( 17 , 27 , 110 ); Dra, Deinococcus radiodurans ( 139 ).
c Alignment score, 30–50.
d BLAST search (tblastn) identified a homologous gene here, but no coding sequence is assigned in the database.
e Alignment score, 50–100.
f In this organism, δ has been recently found, and a τδδ′ complex (without γ) actually acts as a clamp loader ( 16 ).
g These genes are named dnaE in their original databases but are renamed dnaG in this table.