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DciA is an ancestral replicative helicase operator essential for bacterial replication initiation.

ABSTRACT: Delivery of the replicative helicase onto DNA is an essential step in the initiation of replication. In bacteria, DnaC (in Escherichia coli) and DnaI (in Bacillus subtilis) are representative of the two known mechanisms that assist the replicative helicase at this stage. Here, we establish that these two strategies cannot be regarded as prototypical of the bacterial domain since dnaC and dnaI (dna[CI]) are present in only a few bacterial phyla. We show that dna[CI] was domesticated at least seven times through evolution in bacteria and at the expense of one gene, which we rename dciA (dna[CI] antecedent), suggesting that DciA and Dna[CI] share a common function. We validate this hypothesis by establishing in Pseudomonas aeruginosa that DciA possesses the attributes of the replicative helicase-operating proteins associated with replication initiation.

SUBMITTER: Brezellec P

PROVIDER: S-EPMC5109545 | biostudies-literature | 2016 Nov

REPOSITORIES: biostudies-literature

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DciA is an ancestral replicative helicase operator essential for bacterial replication initiation.

Brézellec Pierre P Vallet-Gely Isabelle I Possoz Christophe C Quevillon-Cheruel Sophie S Ferat Jean-Luc JL

Nature communications 20161110

Delivery of the replicative helicase onto DNA is an essential step in the initiation of replication. In bacteria, DnaC (in Escherichia coli) and DnaI (in Bacillus subtilis) are representative of the two known mechanisms that assist the replicative helicase at this stage. Here, we establish that these two strategies cannot be regarded as prototypical of the bacterial domain since dnaC and dnaI (dna[CI]) are present in only a few bacterial phyla. We show that dna[CI] was domesticated at least seve ...[more]

PMID: 27830752

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Project description:A fundamental requirement for life is the replication of an organism's DNA. Studies in Escherichia coli and Bacillus subtilis have set the paradigm for DNA replication in bacteria. During replication initiation in E. coli and B. subtilis, the replicative helicase is loaded onto the DNA at the origin of replication by an ATPase helicase loader. However, most bacteria do not encode homologs to the helicase loaders in E. coli and B. subtilis. Recent work has identified the DciA protein as a predicted helicase operator that may perform a function analogous to the helicase loaders in E. coli and B. subtilis. DciA proteins, which are defined by the presence of a DUF721 domain (termed the DciA domain herein), are conserved in most bacteria but have only been studied in mycobacteria and gammaproteobacteria (Pseudomonas aeruginosa and Vibrio cholerae). Sequences outside the DciA domain in Mycobacterium tuberculosis DciA are essential for protein function but are not conserved in the P. aeruginosa and V. cholerae homologs, raising questions regarding the conservation and evolution of DciA proteins across bacterial phyla. To comprehensively define the DciA protein family, we took a computational evolutionary approach and analyzed the domain architectures and sequence properties of DciA domain-containing proteins across the tree of life. These analyses identified lineage-specific domain architectures among DciA homologs, as well as broadly conserved sequence-structural motifs. The diversity of DciA proteins represents the evolution of helicase operation in bacterial DNA replication and highlights the need for phylum-specific analyses of this fundamental biological process. IMPORTANCE Despite the fundamental importance of DNA replication for life, this process remains understudied in bacteria outside Escherichia coli and Bacillus subtilis. In particular, most bacteria do not encode the helicase-loading proteins that are essential in E. coli and B. subtilis for DNA replication. Instead, most bacteria encode a DciA homolog that likely constitutes the predominant mechanism of helicase operation in bacteria. However, it is still unknown how DciA structure and function compare across diverse phyla that encode DciA proteins. In this study, we performed computational evolutionary analyses to uncover tremendous diversity among DciA homologs. These studies provide a significant advance in our understanding of an essential component of the bacterial DNA replication machinery.

Project description:Replicative helicases play central roles in chromosome duplication and their assembly onto DNA is regulated via initiators and helicase loader proteins. The Escherichia coli replicative helicase DnaB and the helicase loader DnaC form a DnaB6-DnaC6 complex that is required for loading DnaB onto single-stranded DNA. Overexpression of dnaC inhibits replication by promoting continual rebinding of DnaC to DnaB and consequent prevention of helicase translocation. Here we show that overexpression of dnaB also inhibits growth and chromosome duplication. This inhibition is countered by co-overexpression of wild-type DnaC but not of a DnaC mutant that cannot interact with DnaB, indicating that a reduction in DnaB6-DnaC6 concentration is responsible for the phenotypes associated with elevated DnaB concentration. Partial defects in the oriC-specific initiator DnaA and in PriA-specific initiation away from oriC during replication repair sensitise cells to dnaB overexpression. Absence of the accessory replicative helicase Rep, resulting in increased replication blockage and thus increased reinitiation away from oriC, also exacerbates DnaB-induced defects. These findings indicate that elevated levels of helicase perturb replication initiation not only at origins of replication but also during fork repair at other sites on the chromosome. Thus, imbalances in levels of the replicative helicase and helicase loader can inhibit replication both via inhibition of DnaB6-DnaC6 complex formation with excess DnaB, as shown here, and promotion of formation of DnaB6-DnaC6 complexes with excess DnaC [Allen GC, Jr., Kornberg A. Fine balance in the regulation of DnaB helicase by DnaC protein in replication in Escherichia coli. J. Biol. Chem. 1991;266:22096-22101; Skarstad K, Wold S. The speed of the Escherichia coli fork in vivo depends on the DnaB:DnaC ratio. Mol. Microbiol. 1995;17:825-831]. Thus, there are two mechanisms by which an imbalance in the replicative helicase and its associated loader protein can inhibit genome duplication.

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DciA is an ancestral replicative helicase operator essential for bacterial replication initiation.

Publications

DciA is an ancestral replicative helicase operator essential for bacterial replication initiation.

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