Mechanism of Inhibition of Deoxyribonucleic Acid Synthesis in
            <i>Escherichia coli</i>
            by Hydroxyurea

Sinha, Navin K.; Snustad, D. Peter

doi:10.1128/jb.112.3.1321-1334.1972

Cited by 81 publications

(23 citation statements)

References 33 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Repeated trials failed to generate any authentic mutants implying that deletion of both genes is lethal and that S. coelicolor does not possess any other functional RNR systems (indeed, no other genes encoding RNR-like proteins are present in the complete genome sequences of S. coelicolor and Streptomyces avermitilis ). Correlation of this conclusion was obtained from a study of the effect of hydroxyurea -a potent inhibitor of class I RNRs (Sinha and Snustad, 1972;Jordan et al ., 1994) -on the growth of M145 and the nrdB and nrdJ deletion mutant strains. Figure 2 shows that all three strains, M145, M145 D nrdB::apr and M145 D nrdJ::apr , grew equally well on solid medium whereas the M145 D nrdJ::apr strain alone failed to grow in the presence of 10 mM hydroxyurea.…”

Section: Resultsmentioning

confidence: 86%

Alternative oxygen‐dependent and oxygen‐independent ribonucleotide reductases in Streptomyces: cross‐regulation and physiological role in response to oxygen limitation

Borovok

Gorovitz

Yanku

et al. 2004

Molecular Microbiology

View full text Add to dashboard Cite

SummaryRibonucleotide reductases (RNRs) catalyse the conversion of ribonucleotides to deoxyribonucleotides and are essential for de novo DNA synthesis and repair. Streptomyces spp. contain genes coding for two RNRs. We show here that the Streptomyces coelicolor M145 nrdAB genes encoding an oxygendependent class I RNR are co-transcribed with nrdS , which encodes an AraC-like regulatory protein. Likewise, the class II oxygen-independent RNR nrdJ gene forms an operon with a likely regulatory gene, nrdR , which encodes a protein possessing an ATP-cone domain like those present in the allosteric activity site of many class Ia RNRs. Deletions in nrdB and nrdJ had no discernible effect on growth individually, but abolition of both RNR systems, using hydroxyurea to inactivate the class Ia RNR (NrdAB) in the nrdJ deletion mutant, was lethal, establishing that S. coelicolor possesses just two functional RNR systems. The class II RNR (NrdJ) may function to provide a pool of deoxyribonucleotide precursors for DNA repair during oxygen limitation and/or for immediate growth after restoration of oxygen, as the nrdJ mutant was slower in growth recovery than the nrdB mutant or the parent strain. The class Ia and class II RNR genes show complex regulation. The nrdRJ genes were transcribed some five-to sixfold higher than the nrdABS genes in vegetative growth, but when nrdJ was deleted, nrdABS transcription was upregulated by 13-fold. In a reciprocal experiment, deletion of nrdB had little effect on nrdRJ transcription. Deletion of nrdR caused a dramatic increase in transcription of nrdJ and to a less extent nrdABS , whereas disruption of cobN , a gene required for synthesis of coenzyme B12 a cofactor for the class II RNR, caused similar upregulation of transcription of nrdRJ and nrdABS . In contrast, deletion of nrdS had no detectable effect on transcription of either set of RNR genes. These results establish the existence of control mechanisms that sense and regulate overall RNR gene expression.

show abstract

Section: Resultsmentioning

confidence: 86%

Alternative oxygen‐dependent and oxygen‐independent ribonucleotide reductases in Streptomyces: cross‐regulation and physiological role in response to oxygen limitation

Borovok

Gorovitz

Yanku

et al. 2004

Molecular Microbiology

View full text Add to dashboard Cite

show abstract

“…This agrees with the fact that the presence of this transposon eliminates the temperature sensitivity in that mutant. Hydroxyurea, which is a strong inhibitor of the activity of both nrdABand nrdEF-encoded ribonucleotide reductases (Sinha and Snustad, 1972;Jordan et al, 1994b) and stimulates nrdAB transcription (Gibert et al, 1990), increased nrdEF expression in all cases. Wild-type and nrdA ts S. typhimurium cells presented the same behaviour as that obtained with their respective E. coli strains (data not shown).…”

Section: Quantification Of the Nrdef Operon Expressionmentioning

confidence: 96%

Promoter identification and expression analysis of Salmonella typhimurium and Escherichia coli nrdEF operons encoding one of two class I ribonucleotide reductases present in both bacteria

Jordan

Aragall

Gibert

et al. 1996

Molecular Microbiology

View full text Add to dashboard Cite

Salmonella typhimurium and Escherichia coli cells have two different class I ribonucleotide reductases encoded by the nrdEF and nrdAB operons. Despite the presence of one additional ribonucleotide reductase, the nrdAB-encoded enzyme is essential to the aerobic growth of the cell because nrdAB-defective mutants of both species are not viable in the presence of oxygen. Several factors controlling nrdAB gene transcription have been analysed intensively. Nothing is known about the expression of the nrdEF genes. To study this subject, and after cloning of E. coli nrdEF genes and sequencing of their 5' ends, the promoter of this operon has been identified by primer extension in both bacterial species. The +1 position was 691 bp and 692 bp upstream of the translational start points of the nrdE genes of S. typhimurium and E. coli, respectively. Downstream of the +1 position, and before the nrdE gene, two open reading frames (ORFs) of 81 and 136 amino acid residues are present in both bacteria. The synthesis of a polypeptide with a molecular mass of 9 kDa, corresponding to the first of these two ORFs, was observed by using the T7 RNA polymerase expression system. Comparison of the amino acid predicted sequence of this ORF reveals a significant similarity with glutaredoxin proteins. Competitive, reverse-transcription polymerase chain reaction experiments indicate that transcription from the nrdEF promoter normally takes place in wild-type cells. nrdEF transcription is increased by hydroxyurea, which inhibits class I ribonucleotide reductase activity, in both RecA+ and RecA- cells. nrdA(ts) mutants show a higher level of nrdEF transcription than wild-type cells at either the permissive or the restrictive temperature. nrdEF expression was unaffected by changes in DNA supercoiling whether caused by the introduction of either topA::Tn10 and hns::Tn10 mutations or by the inhibition of DNA gyrase with the antibiotic novobiocin. In contrast to the nrdAB genes, the nrdEF operon is not essential to the cells because nrdEF-defective mutants are viable under both aerobic and anaerobic conditions.

show abstract

“…In a similar manner, it is possible that FtsH might degrade unutilized FtsZ if septation arrest is induced by DNA replication inhibition (Huisman et al 1980, 1984, Huisman and D'Ari, 1981. AR5090/pSTD113 cultures were induced for FtsH expression for 60 min, and then treated with the DNA replication inhibitors, hydroxyurea, nalidixic acid, phenethylalcohol or mitomycin C at the final concentrations of 66 mM (Sinha and Snustad 1972) or 20 µg/ml (Goss et al 1965) or 0.4% (Kaneko et al 1977) or 3 µg/ml (Khil and Camerini-Otero 2002), respectively, for further 60 min. Denaturing immunoprecipitation of FtsZ from the whole cell lysates of these cultures grown for a further period of 60 min, followed by pulse-chase, showed that FtsZ was as stable as in the exponentially growing cultures (Fig.…”

Section: Ftsh Does Not Degrade Ftsz Even Under Cell Division Arrestedmentioning

confidence: 99%

Bacterial cell division protein FtsZ is stable against degradation by AAA family protease FtsH in Escherichia coli cells

Srinivasan

Ajitkumar

2007

J. Basic Microbiol.

View full text Add to dashboard Cite

We have found that FtsH protease of Escherichia coli could degrade E. coli cell division protein FtsZ in an ATP- and Zn(2+)-dependent manner in vitro and that the degradation did not show specificity for the N-terminus or C-terminus of FtsZ, like in the case of degradation of its conventional substrate sigma(32) protein. In continuation of these observations, in the present study, we examined whether FtsH would affect the stability and turnover of FtsZ in vivo. We found that FtsZ levels were not elevated in E. coli AR754 (ftsH1 ts) cells at nonpermissive temperature as compared to the levels in an FtsH-active isogenic AR753 strain. Neither did FtsH degrade ectopically expressed FtsZ in AR754 strain nor did ectopic expression of FtsH reduced FtsZ levels in E. coli AR5090 ftsH null strain (ftsH::kan, sfhC21). Pulse chase experiments in AR754 and AR5090 strains showed that there were no compensatory changes in FtsZ turnover, in case FtsZ degradation had occurred. Even under cell division arrested conditions, wherein FtsZ was not required, FtsH protease did not degrade unutilized FtsZ. These experiments demonstrate that either FtsH protease may not have a role in regulating the levels of FtsZ in vivo under the conditions tested or that some cellular component(s) might be stabilising FtsZ against FtsH protease.

show abstract

Mechanism of Inhibition of Deoxyribonucleic Acid Synthesis in Escherichia coli by Hydroxyurea

Cited by 81 publications

References 33 publications

Alternative oxygen‐dependent and oxygen‐independent ribonucleotide reductases in Streptomyces: cross‐regulation and physiological role in response to oxygen limitation

Alternative oxygen‐dependent and oxygen‐independent ribonucleotide reductases in Streptomyces: cross‐regulation and physiological role in response to oxygen limitation

Promoter identification and expression analysis of Salmonella typhimurium and Escherichia coli nrdEF operons encoding one of two class I ribonucleotide reductases present in both bacteria

Bacterial cell division protein FtsZ is stable against degradation by AAA family protease FtsH in Escherichia coli cells

Contact Info

Product

Resources

About