Low Affinity DnaA-ATP Recognition Sites in E. coli oriC Make Non-equivalent and Growth Rate-Dependent Contributions to the Regulated Timing of Chromosome Replication

Rao, Prassanna; Rozgaja, Tania A.; Alqahtani, Abdulaziz; Grimwade, Julia E.; Leonard, Alan C.

doi:10.3389/fmicb.2018.01673

Cited by 8 publications

(15 citation statements)

References 66 publications

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“…IHF was discovered by investigators of the life cycle of bacteriophage lambda because this protein is essential for the integration into and excision from the E. coli chromosome through site-specific recombination at the att l site [38]. Both proteins also play important structural roles in the initiation of chromosome replication [39,40]. These structural observations predisposed the field to consider NAPs from an architectural perspective first, with discoveries of their involvement in transcription control following later.…”

Section: When Is a Transcription Factor A Nap?mentioning

confidence: 99%

When is a transcription factor a NAP?

Dorman

Schumacher

Bush

et al. 2020

Current Opinion in Microbiology

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Proteins that regulate transcription often also play an architectural role in the genome. Thus, it has been difficult to define with precision the distinctions between transcription factors and nucleoid-associated proteins (NAPs). Anachronistic descriptions of NAPs as 'histone-like' implied an organizational function in a bacterial chromatin-like complex. Definitions based on protein abundance, regulatory mechanisms, target gene number, or the features of their DNAbinding sites are insufficient as marks of distinction, and trying to distinguish transcription factors and NAPs based on their ranking within regulatory hierarchies or positions in genecontrol networks is also unsatisfactory. The terms 'transcription factor' and 'NAP' are ad hoc operational definitions with each protein lying along a spectrum of structural and functional features extending from highly specific actors with few gene targets to those with a pervasive influence on the transcriptome. The Streptomyces BldC protein is used to illustrate these issues.

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Section: When Is a Transcription Factor A Nap?mentioning

confidence: 99%

When is a transcription factor a NAP?

Dorman

Schumacher

Bush

et al. 2020

Current Opinion in Microbiology

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“…As DnaA-ATP levels increase during the cell cycle, progressive DnaA occupation of the right array of sites displaces Fis (Ryan et al, 2004), allowing IHF to bind, resulting in a DNA bend that places R1 sufficiently close to R5M to nucleate filling of oriC ’s left side low affinity sites (Rozgaja et al, 2011). By acting as a temporary partition between the left and right halves of oriC (Gille et al, 1991), Fis is able to delay initiation until the total number of DnaA molecules in the cell exceeds that needed for initiation of a single oriC copy; thus, when Fis is finally displaced, all origins in the cell can complete orisome assembly and initiate synchronously (Ryan et al, 2004; Rao et al, 2018). In this way, Fis becomes the primary regulator of initiation timing under rapid growth conditions (Flåtten and Skarstad, 2013).…”

Section: Ordered Orisome Assemblymentioning

confidence: 99%

Changing Perspectives on the Role of DnaA-ATP in Orisome Function and Timing Regulation

et al. 2019

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Bacteria, like all cells, must precisely duplicate their genomes before they divide. Regulation of this critical process focuses on forming a pre-replicative nucleoprotein complex, termed the orisome. Orisomes perform two essential mechanical tasks that configure the unique chromosomal replication origin, oriC to start a new round of chromosome replication: (1) unwinding origin DNA and (2) assisting with loading of the replicative DNA helicase on exposed single strands. In Escherichia coli , a necessary orisome component is the ATP-bound form of the bacterial initiator protein, DnaA. DnaA-ATP differs from DnaA-ADP in its ability to oligomerize into helical filaments, and in its ability to access a subset of low affinity recognition sites in the E. coli replication origin. The helical filaments have been proposed to play a role in both of the key mechanical tasks, but recent studies raise new questions about whether they are mandatory for orisome activity. It was recently shown that a version of E. coli oriC ( oriC allADP ), whose multiple low affinity DnaA recognition sites bind DnaA-ATP and DnaA-ADP similarly, was fully occupied and unwound by DnaA-ADP in vitro , and in vivo suppressed the lethality of DnaA mutants defective in ATP binding and ATP-specific oligomerization. However, despite their functional equivalency, orisomes assembled on oriC allADP were unable to trigger chromosome replication at the correct cell cycle time and displayed a hyper-initiation phenotype. Here we present a new perspective on DnaA-ATP, and suggest that in E. coli , DnaA-ATP is not required for mechanical functions, but rather is needed for site recognition and occupation, so that initiation timing is coupled to DnaA-ATP levels. We also discuss how other bacterial types may utilize DnaA-ATP and DnaA-ADP, and whether the high diversity of replication origins in the bacterial world reflects different regulatory strategies for how DnaA-ATP is used to control orisome assembly.

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“…Fis is a prominent member of the family of NAPs in Gram-negative bacteria (13,14) that alters the transcription of hundreds of genes, directly or indirectly and positively or negatively (15)(16)(17)(18)(19), and contributes architecturally to site-specific recombination systems (20)(21)(22), chromosome replication initiation (23)(24)(25)(26), transposon activity (27,28), and bacteriophage life cycles (22,27,29,30). Fis is not essential, despite its pervasive influence on cell biology, but it enhances the fitness of a wild-type (WT) bacterium when competing with an otherwise isogenic fis knockout mutant (31).…”

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confidence: 99%

Network Rewiring: Physiological Consequences of Reciprocally Exchanging the Physical Locations and Growth-Phase-Dependent Expression Patterns of the Salmonella fis and dps Genes

Bogue

Mogre

Beckett

et al. 2020

mBio

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The Fis nucleoid-associated protein controls the expression of a large and diverse regulon of genes in Gram-negative bacteria. Fis production is normally maximal in bacteria during the early exponential phase of batch culture growth, becoming almost undetectable by the onset of stationary phase. We tested the effect on the Fis regulatory network in Salmonella of moving the complete fis gene from its usual location near the origin of chromosomal replication to the position normally occupied by the dps gene in the right macrodomain of the chromosome, and vice versa, creating the gene exchange (GX) strain. In a parallel experiment, we tested the effect of rewiring the Fis regulatory network by placing the fis open reading frame under the control of the stationary-phase-activated dps promoter at the dps genetic location within the right macrodomain, and vice versa, creating the open reading frame exchange (OX) strain. Chromatin immunoprecipitation sequencing (ChIP-seq) was used to measure global Fis protein binding levels and to determine gene expression patterns. Strain GX showed few changes compared with the wild type, although we did detect increased Fis binding at Ter, accompanied by reduced binding at Ori. Strain OX displayed a more pronounced version of this distorted Fis protein-binding pattern together with numerous alterations in the expression of genes in the Fis regulon. OX, but not GX, had a reduced ability to infect cultured mammalian cells. These findings illustrate the inherent robustness of the Fis regulatory network with respect to the effects of rewiring based on gene repositioning alone and emphasize the importance of fis expression signals in phenotypic determination. IMPORTANCE We assessed the impact on Salmonella physiology of reciprocally translocating the genes encoding the Fis and Dps nucleoid-associated proteins (NAPs) and of inverting their growth-phase production patterns such that Fis was produced in stationary phase (like Dps) and Dps was produced in exponential phase (like Fis). Changes to peak binding of Fis were detected by ChIP-seq on the chromosome, as were widespread impacts on the transcriptome, especially when Fis production mimicked Dps production. Virulence gene expression and the expression of a virulence phenotype were altered. Overall, these radical changes to NAP gene expression were well tolerated, revealing the robust and well-buffered nature of global gene regulation networks in the bacterium.

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Low Affinity DnaA-ATP Recognition Sites in E. coli oriC Make Non-equivalent and Growth Rate-Dependent Contributions to the Regulated Timing of Chromosome Replication

Cited by 8 publications

References 66 publications

When is a transcription factor a NAP?

When is a transcription factor a NAP?

Changing Perspectives on the Role of DnaA-ATP in Orisome Function and Timing Regulation

Network Rewiring: Physiological Consequences of Reciprocally Exchanging the Physical Locations and Growth-Phase-Dependent Expression Patterns of the Salmonella fis and dps Genes

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