The origin of replication, oriC, and the dnaA protein are dispensable in stable DNA replication (sdrA) mutants of Escherichia coli K-12.

Kogoma, Tokio; Meyenburg, Kaspar von

doi:10.1002/j.1460-2075.1983.tb01445.x

Cited by 118 publications

(85 citation statements)

References 27 publications

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“…In rnhA224 mutant cells (AQ10033), an alternative replication system, constitutive stable DNA replication (cSDR) is activated and thus oriC defects are suppressed (33). Using this strain, we successfully transferred the triple mutant oriC onto the chromosome.…”

Section: Resultsmentioning

confidence: 99%

The absence of effect of gid or mioC transcription on the initiation of chromosomal replication in Escherichia coli

Bates

Boye²,

Asai

et al. 1997

Proc. Natl. Acad. Sci. U.S.A.

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Despite the widely accepted view that transcription of gid and mioC is required for efficient initiation of cloned oriC, we show that these transcriptions have very little effect on initiation of chromosome replication at wild-type chromosomal oriC. Furthermore, neither gid nor mioC transcription is required in cells deficient in the histone-like proteins Fis or IHF. However, oriC that is sufficiently impaired for initiation by deletion of DnaA box R4 requires transcription of at least one of these genes. We conclude that transcription of mioC and especially gid is needed to activate oriC only under suboptimal conditions. We suggest that either the rifampicin-sensitive step of initiation is some other transcription occurring from promoter(s) within oriC, or the original inference of transcriptional activation derived from the rifampicin experiments is incorrect.Initiation of replication of the Escherichia coli chromosome occurs at a unique site, oriC. Initiation depends primarily on DnaA protein, which binds to five 9-bp repeats within oriC and facilitates duplex melting at three AϩT-rich 13-mers (see Fig. 1). DnaB helicase subsequently enters the opened duplex and leads to priming and chain elongation reactions resulting in bidirectional replication. oriC is a complex regulatory region containing recognition sites for many positive and negative regulatory proteins, including Fis, IHF, SeqA, and IciA, some or all of which are thought to help precisely regulate initiations occurring at oriC (see ref. 1 for review).Early physiology experiments by Lark (2) and Messer (3) first suggested that RNA polymerase (RNAP) is somehow involved in initiation at oriC. This was inferred from findings that rifampicin (an inhibitor of RNAP) inhibits a new round of DNA synthesis at a time when protein synthesis is no longer required. Genetic evidence as well suggests a role of RNAP in initiation. Mutations in rpoB and rpoC, which encode the ␤ and ␤Ј subunits of RNAP, respectively, have been shown to increase copy numbers of both the chromosome and chimeric plasmids (oriC plasmids) carrying both an oriC site and a ColE1-type replication origin (4, 5). Further, specific rpoB mutations have been shown to suppress the temperature sensitivity phenotype of certain dnaA(Ts) mutations (6). Despite the evidence suggesting an involvement of RNAP, a specific transcription event required for initiation has not been identified.Of promoters possibly involved in transcriptional activation of initiation, one likely candidate is the promoter of gidAB. The gid promoter (P gid ) is situated just counterclockwise of oriC ( Fig. 1) and transcription proceeds leftward away from oriC. oriC plasmids in which the gid promoter has been inactivated exhibit decreased transformation efficiency and stability as well as decreased replication in vitro (7,8). The twin-domain model of Liu and Wang (9) predicts that an actively transcribing RNAP generates local domains of increased negative supercoiling behind it and positive supercoiling in front of it. It...

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Section: Resultsmentioning

confidence: 99%

The absence of effect of gid or mioC transcription on the initiation of chromosomal replication in Escherichia coli

Bates

Boye²,

Asai

et al. 1997

Proc. Natl. Acad. Sci. U.S.A.

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“…The presence of the oriCdel-1071 mutation in AoriC of the dnaA::TnlO mutation inhibited the transformation with strains was verified by Southern blot hybridization as described pOC81. pFHC539 carrying dnaA+ restored the ability to previously (15). pOC81 (4) and pFHC359 and pDKT143 (14) support pOC81 replication to the dnaA::TnJO strain but were described previously.…”

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confidence: 82%

“…In RNase HI-deficient (rnhA) mutants, initiation can occur at multiple sites (oriKs), which are normally repressed in wild-type cells (see reference 11 for a review). Thus, as long as oriK sites are active, the oriC site and DnaA protein are dispensable in rnhA mutants (15). The replication from oriK sites in the absence of RNase HI, designated constitutive stable DNA replication, can be initiated in the presence of chloramphenicol, an inhibitor of protein synthesis, whereas continuous initiation from oriC strictly requires protein synthesis (11).…”

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Absence of a direct role for RNase HI in initiation of DNA replication at the oriC site on the Escherichia coli chromosome

Hong

Kogoma

1993

J Bacteriol

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On the basis of the experiments carried out with rnhA224 mutants, we previously concluded that RNase HI is not essential for initiation of Escherichia coli chromosome replication at oriC (T. Kogoma, N.L. Subia, and K. von Meyenburg, Mol. Gen. Genet. 200:103-109, 1985). In light of the recent finding that rnhA224 is a UGA nonsense mutation which can be leaky in certain genetic backgrounds, we reexamined this conclusion with the use of rnhA339 (Null)::cat mutants. The possibility that recB+ is required for initiation at the alternative origins (oriKs) of replication in rnhA mutants was also tested. The results clearly indicated that RNase HI is not essential for oriC initiation and that recB+ is not required for initiation at oriK sites.

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“…For example, DnaA-dependent initiation of DNA replication at the Escherichia coli oriC replication origin can be overcome in the absence of RNase H1 (28,29). As a consequence, rnhA mutants can survive complete inactivation of oriC by transcriptiondependent activation of so-called oriK sites (30,31), although candidate oriK sites have been identified only recently (32). Additional evidence for R-loop-primed replication was given by the observation that rnhA mutants are prone to an increase in mutation and DNA amplification events if origin activity is suppressed.…”

mentioning

confidence: 99%

Role for RNA:DNA hybrids in origin-independent replication priming in a eukaryotic system

Stuckey

García‐Rodríguez

Aguilera

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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DNA replication initiates at defined replication origins along eukaryotic chromosomes, ensuring complete genome duplication within a single S-phase. A key feature of replication origins is their ability to control the onset of DNA synthesis mediated by DNA polymerase-α and its intrinsic RNA primase activity. Here, we describe a novel origin-independent replication process that is mediated by transcription. RNA polymerase I transcription constraints lead to persistent RNA:DNA hybrids (R-loops) that prime replication in the ribosomal DNA locus. Our results suggest that eukaryotic genomes have developed tools to prevent R-loop-mediated replication events that potentially contribute to copy number variation, particularly relevant to carcinogenesis.D uring transcription, DNA acts as a template for the synthesis of the nascent RNA. RNA synthesis is accompanied by the generation of positive and negative DNA supercoiling in front of and behind the transcription machinery, respectively (1). Unwinding of the DNA double helix by negative supercoiling may allow the RNA to hybridize to its DNA template behind the elongating RNA polymerase, leading to R-loops (2). Other elements that could potentiate R-loop accumulation include RNA: DNA hybrid-facilitating DNA sequences, such as G-quadruplex structures (3) or nicks in the nontemplate DNA strand (4).Eukaryotic cells need to control R-loop formation to avoid replication impairment, genome instability, and life span shortening mediated by such intermediates (5-10; reviewed in ref. 11). To do so, cells catalyze the relaxation of supercoiled DNA by type I topoisomerases (12-15), thus preventing replication fork reversal (16), DNA overwinding with the potential to block replication fork progression (17), DNA unwinding (18), or R-loop-mediated blocks of ribosomal RNA synthesis (19). Other enzymatic activities involved in R-loop processing include ribonuclease H (RNase H) activities, DNA-RNA helicases, such as Sen1/senataxin (20, 21), or Ataxin-2 RNA-binding protein Pbp1 (10). The ribonuclease activity of Saccharomyces cerevisiae RNases H1 and H2 specifically cleaves the RNA moiety of the RNA:DNA hybrid structure (22), whereas RNase H2 and topoisomerase 1 (Top1) can also process ribonucleotides in duplex DNA (23,24).Notably, R-loops are required to initiate mitochondrial DNA replication (25) and pioneering studies connected R-loops to origin-independent replication in prokaryotic systems (26,27). For example, DnaA-dependent initiation of DNA replication at the Escherichia coli oriC replication origin can be overcome in the absence of RNase H1 (28, 29). As a consequence, rnhA mutants can survive complete inactivation of oriC by transcriptiondependent activation of so-called oriK sites (30, 31), although candidate oriK sites have been identified only recently (32). Additional evidence for R-loop-primed replication was given by the observation that rnhA mutants are prone to an increase in mutation and DNA amplification events if origin activity is suppressed. These events required remov...

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The origin of replication, oriC, and the dnaA protein are dispensable in stable DNA replication (sdrA) mutants of Escherichia coli K-12.

Cited by 118 publications

References 27 publications

The absence of effect of gid or mioC transcription on the initiation of chromosomal replication in Escherichia coli

The absence of effect of gid or mioC transcription on the initiation of chromosomal replication in Escherichia coli

Absence of a direct role for RNase HI in initiation of DNA replication at the oriC site on the Escherichia coli chromosome

Role for RNA:DNA hybrids in origin-independent replication priming in a eukaryotic system

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