Replication of the Bacillus subtilis Chromosome
Philippe Noirot, Patrice Polard and Marie-Françoise Noirot-Gros
from Bacillus: Cellular and Molecular Biology Download flyer
Eubacteria have evolved multicomponent protein machines, termed replisomes, that duplicate their chromosomes rapidly and accurately. Extensive studies in the model bacteria Escherichia coli and Bacillus subtilis have revealed that in addition to the replication core machinery, other proteins are necessary to form a functional replication fork. Specific subsets of proteins mediate a) the assembly of the replisome at the chromosomal origin of replication (initiation), b) the progression of the replication forks along the chromosome (elongation) and their maintenance by providing solutions for replication restart, which are adapted to possible 'roadblocks' encountered on the DNA template, and c) promote the physiological arrest of replication when chromosome duplication is completed (termination). Within the cell, DNA replication takes place within a factory positioned at the cell centre. This review summarises the recent knowledge about chromosomal replication in Bacillus subtilis and related Gram-positive bacteria. It is focused on the events governing the assembly and the fate of the replication fork, describes protein networks connected with the replisome, and emphasises several novel aspects of DNA replication in this group of bacteria. Read more ....
Bacillus subtilis duplicates its single circular chromosome by initiating DNA replication at a single locus, the origin (oriC). Replication proceeds bidirectionally and two replication forks progress in the clockwise and counterclockwise directions along the chromosome halves (Lemon et al., 2002). Chromosome replication is completed when the forks reach the terminus region, which is positioned opposite to the origin on the chromosome map, and contains several short DNA sequences (Ter sites) that promote replication arrest (Duggin and Wake, 2002). Specific proteins mediate all the steps in DNA replication. The comparison between the sets of proteins involved in chromosomal DNA replication in B. subtilis and in Escherichia coli reveals both similarities and differences. Although the basic components promoting initiation, elongation, and termination of replication are well conserved, some important differences can be found (such as one bacterium missing proteins essential in the other). These differences underline the diversity in the mechanisms and strategies that various bacterial species have adopted to carry out the duplication of their genomes.
The level of detail at which DNA replication is understood in B. subtilis is second only to E. coli among prokaryotes. The comparison between both organisms reveals similarities in the overall architecture of the replisome, in the ways DNA replication is initiated at the chromosomal origin and reinitiated at arrested replication forks, and how this results in a spatially and temporally controlled loading of the replicative helicase onto the single-stranded lagging-strand template. The comparison also reveals many striking differences that appear to be highly conserved among Gram-positive bacteria. The biological significance of these differences extends far beyond the expected diversity in the molecular details, and will likely be the focus of the research in the next few years. B. subtilis has two distinct DNA polymerases essential for the elongation of DNA replication that are associated with the replication factory. However, the second polymerase DnaE displays a low fidelity of synthesis and is capable of lesion bypass. What is the function of DnaE in the cell that could reconcile these properties? Another major difference is the involvement of the B. subtilis primosomal proteins DnaB, DnaD and DnaI in both the DnaA-mediated initiation at the origin, and the PriA-mediated replication restart. The DnaB, DnaD and DnaI proteins have DNA binding and remodeling activities, act in helicase loading, and are the targets of mechanisms that regulate replication. The DnaA-dependent initiation is negatively regulated by YabA. How do these regulatory mechanisms distinguish between the initiation at oriC that occurs only once per cell cycle and the replication restart that is needed throughout chromosomal replication? Finally, the B. subtilis DNA polymerase I processes the Okazaki fragments, participates in the repair of DNA damage and plasmid replication, and assists in translesion synthesis mediated by the Y-polymerases. Pol I lacks an exonuclease proofreading activity, in keeping with its role in TLS, but in contradiction with the rather extensive error-free DNA synthesis required for DNA repair and plasmid replication. Does B. subtilis Pol I mediate DNA synthesis in vivo, or does it fulfill a structural role, possibly through its interactions with PolC and DnaE polymerases?
B. subtilis is an organism of choice for cellular biology studies, in which the replication factory has been discovered, and where the subcellular localization of most replication proteins is known. A major challenge is to understand how such a replication factory is constructed, maintained, and disassembled throughout the cell cycle. In combination, the subcellular localization of proteins, the transcriptional responses to perturbations of replication, and the interaction network of replication proteins will help to identify the proteins that act to coordinate DNA replication with other cellular processes. Read more ....