Plasmid replication initiator interactions with origin 13-mers and polymerase subunits contribute to strand-specific replisome assembly

Wawrzycka, Aleksandra; Gross, Marta H; Wasaznik, Anna; Konieczny, Igor

doi:10.1073/pnas.1504926112

Cited by 14 publications

(14 citation statements)

References 72 publications

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“…Then, the δ subunit of clamp loader opens the β-clamp and clamp loader positions it onto primer-template junction, as it is thought to occur during replisome assembly at E. coli oriC . This model is consistent with the results of the in vitro DNA replication experiments performed with the use of ssDNA, containing sequence of RK2 plasmid oriV (Wawrzycka et al, 2015 ). It was demonstrated that TrfA interacts with specific strand of ssDNA of DUE, i.e., the bottom strand, which serves as the site for replisome assembly (Wegrzyn et al, 2014 ; Wawrzycka et al, 2015 ).…”

Section: Replisome Assemblysupporting

confidence: 90%

“…This model is consistent with the results of the in vitro DNA replication experiments performed with the use of ssDNA, containing sequence of RK2 plasmid oriV (Wawrzycka et al, 2015 ). It was demonstrated that TrfA interacts with specific strand of ssDNA of DUE, i.e., the bottom strand, which serves as the site for replisome assembly (Wegrzyn et al, 2014 ; Wawrzycka et al, 2015 ). It can be further speculated that TrfA may assist the clamp loader in recognition of the 3′ end of primer-template junction within the oriV .…”

Section: Replisome Assemblysupporting

confidence: 90%

“…The TrfA ΔLF mutant facilitated the determination of biological relevance of this interaction. The complex of TrfA and β-clamp was found to be the key feature for replisome assembly and thereby for oriV- dependent DNA replication of both supercoiled dsDNA plasmid and ssDNA plasmid in vitro , albeit the clamp loader complex is still crucial (Wawrzycka et al, 2015 ). Hence, the question arises—how do the Rep and clamp loader cooperate to load the β-clamp at plasmid origin ?…”

Section: Replisome Assemblymentioning

confidence: 99%

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Replisome Assembly at Bacterial Chromosomes and Iteron Plasmids

Gross

Uciechowska

Konieczny

2016

Front. Mol. Biosci.

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The proper initiation and occurrence of DNA synthesis depends on the formation and rearrangements of nucleoprotein complexes within the origin of DNA replication. In this review article, we present the current knowledge on the molecular mechanism of replication complex assembly at the origin of bacterial chromosome and plasmid replicon containing direct repeats (iterons) within the origin sequence. We describe recent findings on chromosomal and plasmid replication initiators, DnaA and Rep proteins, respectively, and their sequence-specific interactions with double- and single-stranded DNA. Also, we discuss the current understanding of the activities of DnaA and Rep proteins required for replisome assembly that is fundamental to the duplication and stability of genetic information in bacterial cells.

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Section: Replisome Assemblysupporting

confidence: 90%

Section: Replisome Assemblysupporting

confidence: 90%

Section: Replisome Assemblymentioning

confidence: 99%

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Replisome Assembly at Bacterial Chromosomes and Iteron Plasmids

Gross

Uciechowska

Konieczny

2016

Front. Mol. Biosci.

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“…Binding of Lon WT, LonK371E/K376E/R379E, and LonR306E/K308E/K310E/K311E to TrfA was analyzed by ELISA as described previously (66) with the following modifications. 200 nM TrfA and 1% (w/v) BSA solution (negative control) were immobilized on the 96-well ELISA plate (Costar) for 90 min.…”

Section: Elisamentioning

confidence: 99%

Defining the crucial domain and amino acid residues in bacterial Lon protease for DNA binding and processing of DNA-interacting substrates

Karlowicz

Wegrzyn

Gross

et al. 2017

Journal of Biological Chemistry

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Lon protease previously has been shown to interact with DNA, but the role of this interaction for Lon proteolytic activity has not been characterized. In this study, we used truncated Lon constructs, bioinformatics analysis, and site-directed mutagenesis to identify Lon domains and residues crucial for Lon binding with DNA and effects on Lon proteolytic activity. We found that deletion of Lon's ATPase domain abrogated interactions with DNA. Substitution of positively charged amino acids in this domain in full-length Lon with residues conferring a net negative charge disrupted binding of Lon to DNA. These changes also affected the degradation of nucleic acid-binding protein substrates of Lon, intracellular localization of Lon, and cell morphology. tests revealed that Lon-DNA interactions are essential for Lon activity in cell division control. In summary, we demonstrate that the ability of Lon to bind DNA is determined by its ATPase domain, that this binding is required for processing protein substrates in nucleoprotein complexes, and that Lon may help regulate DNA replication in response to growth conditions.

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“…During initiation of replication, the initiator proteins of iteron‐containing plasmids like those of the IncP‐1 group need to interact with host proteins such as DnaA, DnaB, sliding clamp and protein chaperones (DnaKJ/GrpE, ClpB, ClpX, ClpA) (Konieczny et al ., ; Konieczny and Helinski, ; Wickner et al ., ; Wickner et al ., ; Lu et al ., ; Konieczny and Liberek, ; Zhong et al ., ; Kongsuwan et al ., ; Wawrzycka et al ., ). The mutations that occurred in the trfA1 gene during evolution of pMS0506 in S. oneidensis were concentrated in the region coding for the predicted second helix within the TrfA1 N‐terminus region (Hughes et al ., ).…”

Section: Introductionmentioning

confidence: 97%

Evolved plasmid‐host interactions reduce plasmid interference cost

Yano

Wegrzyn

Loftie-Eaton

et al. 2016

Molecular Microbiology

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SUMMARY Antibiotic selection drives adaptation of antibiotic resistance plasmids to new bacterial hosts, but the molecular mechanisms are still poorly understood. We previously showed that a broad-host-range plasmid was poorly maintained in Shewanella oneidensis, but rapidly adapted through mutations in the replication initiation gene trfA1. Here we examined if these mutations reduced the fitness cost of TrfA1, and whether this was due to changes in interaction with the host’s DNA helicase DnaB. The strains expressing evolved TrfA1 variants showed a higher growth rate than those expressing ancestral TrfA1. The evolved TrfA1 variants showed a lower affinity to the helicase than ancestral TrfA1 and were no longer able to activate the helicase at the oriV without host DnaA. Moreover, persistence of the ancestral plasmid was increased upon overexpression of DnaB. Finally, the evolved TrfA1 variants generated higher plasmid copy numbers than ancestral TrfA1. The findings suggest that ancestral plasmid instability can at least partly be explained by titration of DnaB by TrfA1. Thus under antibiotic selection resistance plasmids can adapt to a novel bacterial host through partial loss of function mutations that simultaneously increase plasmid copy number and decrease unfavorably high affinity to one of the hosts’ essential proteins.

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Plasmid replication initiator interactions with origin 13-mers and polymerase subunits contribute to strand-specific replisome assembly

Cited by 14 publications

References 72 publications

Replisome Assembly at Bacterial Chromosomes and Iteron Plasmids

Replisome Assembly at Bacterial Chromosomes and Iteron Plasmids

Defining the crucial domain and amino acid residues in bacterial Lon protease for DNA binding and processing of DNA-interacting substrates

Evolved plasmid‐host interactions reduce plasmid interference cost

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