Abstract
During the cell division cycle of all bacteria, DNA-protein complexes termed orisomes trigger the onset of chromosome duplication. Orisome assembly is both staged and stringently regulated to ensure that DNA synthesis begins at a precise time and only once at each origin per cycle. Orisomes comprise multiple copies of the initiator protein DnaA, which oligomerizes after interacting with specifically positioned recognition sites in the unique chromosomal replication origin, oriC. Since DnaA is highly conserved, it is logical to expect that all bacterial orisomes will share fundamental attributes. Indeed, although mechanistic details remain to be determined, all bacterial orisomes are capable of unwinding oriC DNA and assisting with loading of DNA helicase onto the single-strands. However, comparative analysis of oriCs reveals that the arrangement and number of DnaA recognition sites is surprisingly variable among bacterial types, suggesting there are many paths to produce functional orisome complexes. Fundamental questions exist about why these different paths exist and which features of orisomes must be shared among diverse bacterial types. In this review we present the current understanding of orisome assembly and function in Escherichia coli and compare the replication origins among the related members of the Gammaproteobacteria. From this information we propose that the diversity in orisome assembly reflects both the requirement to regulate the conformation of origin DNA as well as to provide an appropriate cell cycle timing mechanism that reflects the lifestyle of the bacteria. We suggest that identification of shared steps in orisome assembly may reveal particularly good targets for new antibiotics.
Highlights
As the commitment step for proliferation, initiating new rounds of chromosomal DNA synthesis is arguably the paramount event in the life of a bacterial cell. It is a precarious step, which must rely on sophisticated regulatory mechanisms to ensure that new replication forks are established with sufficient time and number to provide every daughter cell with at least one complete genome copy, regardless of cellular growth rate. Many of these regulatory mechanisms are focused on orisomes, the large multimeric complexes of the bacterial initiator protein, DnaA, that assemble along the unique chromosomal replication origin, oriC
Since the spacing between high and low affinity sites in oriC varies among bacterial types, and since domain I interactions play a key role in cooperative binding at oriC (Miller et al, 2009), this stringency in spacing implies that the length of the flexible linker region in DnaA’s domain II may contribute to the efficiency of orisome assembly
Extension of DnaA from R4 is accomplished, due to close spacing (3 bp) of R4 and C1 (Rozgaja et al, 2011), but there is a 45 bp gap between R1 and the nearest low affinity recognition site, R5M, and there is evidence that DnaA oligomers nucleated at R1 do not extend into this region (Rozgaja et al, 2011). To span this 45 bp space, E. coli oriC DNA is bent to bring R1 and R5 into proximity, and the DnaA oligomer that grows by progressive binding of DnaA monomers to the left array of low affinity sites is most likely nucleated by cross-strand DnaA–DnaA interactions via domain I (Figure 3)
Summary
As the commitment step for proliferation, initiating new rounds of chromosomal DNA synthesis is arguably the paramount event in the life of a bacterial cell. Since the spacing between high and low affinity sites in oriC varies among bacterial types, and since domain I interactions play a key role in cooperative binding at oriC (Miller et al, 2009), this stringency in spacing implies that the length of the flexible linker region in DnaA’s domain II may contribute to the efficiency of orisome assembly.
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