Mutation and killing of Escherichia coli expressing a cloned Bacillus subtilis gene whose product alters DNA conformation

Setlow, Jane K.; Randesi, Matthew; Adams, Joel C.; Setlow, Barbara; Setlow, Peter

doi:10.1128/jb.174.9.2943-2950.1992

Cited by 18 publications

(11 citation statements)

References 35 publications

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“…Evidence consistent with the existence of distinct domains in the membrane of growing cells of B. subtilis has been presented (49). It was also reported that the inner membrane of B. megaterium spores could be resolved into approximately equivalent amounts of two fractions of similar lipid composition, but markedly different protein content and composition (50). In contrast, the inner membrane of germinated spores gave rise to only a single fraction that had a protein content and composition intermediate between those of the two fractions from the dormant spore's inner membrane.…”

Section: Resultsmentioning

confidence: 61%

Lipids in the inner membrane of dormant spores of Bacillus species are largely immobile

Cowan

Olivastro²,

Koppel³

et al. 2004

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

175

185

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Bacterial spores of various Bacillus species are impermeable or exhibit low permeability to many compounds that readily penetrate germinated spores, including methylamine. We now show that a lipid probe in the inner membrane of dormant spores of Bacillus megaterium and Bacillus subtilis is largely immobile, as measured by fluorescence redistribution after photobleaching, but becomes free to diffuse laterally upon spore germination. The lipid immobility in and the slow permeation of methylamine through the inner membrane of dormant spores may be due to a significant (1.3-to 1.6-fold) apparent reduction of the membrane surface area in the dormant spore relative to that in the germinated spore, but is not due to the dormant spore's high levels of dipicolinic acid and divalent cations.D ormant bacterial spores of various Bacillus species are extremely resistant to chemical agents that readily kill germinated spores or growing cells (1, 2). One factor important in dormant spore resistance to such chemicals appears to be their slow permeation into the spore core or protoplast. A variety of data indicate that it is the spore's inner membrane that is the major permeability barrier restricting the passage of small molecules into the spore core (3-7), although the lipid composition of the inner membrane exhibits no anomalies that might explain its unusual properties (8)(9)(10)(11)(12). In addition, the dormant spore's inner membrane has the potential to expand significantly, because electron microscopy indicates that the volume this membrane encompasses (the spore core): (i) decreases as much as 2-fold late in sporulation (13); and (ii) increases up to 2-fold in the first minute of spore germination, when the spore's large peptidoglycan cortex is degraded and the germ cell wall expands (13)(14)(15). This increase in core volume during spore germination takes place without new membrane synthesis, because ATP production is not required, and restores relatively normal permeability to the germinated spore's plasma membrane (4,5,16,17).Whereas dyes that stain growing cells, including fluorescent lipid probes, do not stain dormant spores (B.S. and P.S., unpublished results) as expected (15, 18), lipid probes can be incorporated into the developing spore's membranes during sporulation (19). Given the latter finding, we decided to add fluorescent lipid probes during sporulation to prepare spores with labeled membranes, and then use the technique of fluorescence redistribution after photobleaching (FRAP) to directly analyze the fluidity of the spore's inner membrane, because this technique has been used successfully in analyzing the fluidity and organization of molecules in a number of other systems, including spores of Bacillus species (20-25). MethodsStrains and Spore and Cell Preparation. The iosgenic Bacillus subtilis strains used in this work are derivatives of strain 168 and were: PS832, a prototrophic derivative of strain 168; PS3207, cwlD::cam, in which the cwlD gene responsible for production of muramic acid-␦-lactam in ...

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Section: Resultsmentioning

confidence: 61%

Lipids in the inner membrane of dormant spores of Bacillus species are largely immobile

Cowan

Olivastro²,

Koppel³

et al. 2004

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

175

185

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“…8, lanes 6, 7, and 8). Control experiments showed that the binding specificity of SspC't to pUC19 at pH 5.0 was essentially the same as at pH 7.0, as the pattern of DNase II-protected bands from pUC19 with saturating and subsaturating levels of SspCw`was identical at both pHs (27).…”

Section: Resultsmentioning

confidence: 99%

“…The question then becomes why an abasic site generated by a depurination event in spores would be cleaved rapidly. Control experiments have shown that at most 20% of abasic sites undergo strand cleavage during the extraction and nucleic acid purification procedure used (27). Consequently, if the singlestrand breaks are due to depurination, cleavage at the abasic sites must have taken place in vivo.…”

Section: Discussionmentioning

confidence: 99%

Prevention of DNA damage in spores and in vitro by small, acid-soluble proteins from Bacillus species

1993

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The DNA in dormant spores of Bacillus species is saturated with a group of nonspecific DNA-binding proteins, termed oa/13-type small, acid-soluble spore proteins (SASP). These proteins alter DNA structure in vivo and in vitro, providing spore resistance to UV light. In addition, heat treatments (e.g., 850C for 30 min) which give little killing of wild-type spores of B. subtilis kill >99%o of spores which lack most a/13-type SASP (termed a-1-spores). Similar large differences in survival of wild-type and a-13-spores were found at 90, 80, 65, 22, and 10'C. After heat treatment (850C for 30 min) or prolonged storage (220C for 6 months) that gave >99%o killing of at-13-spores, 10 to 20%o of the survivors contained auxotrophic or asporogenous mutations.However, at-A-spores heated for 30 min at 850C released no more dipicolinic acid than similarly heated wild-type spores (<20%o of the total dipicolinic acid) and triggered germination normally. In contrast, after a heat treatment (930C for 30 min) that gave .99%6 killing of wild-type spores, <1% of the survivors had acquired new obvious mutations, >85% of the spore's dipicolinic acid had been released, and <1% of the surviving spores could initiate spore germination. Analysis of DNA extracted from heated (850C, 30 min) and unheated wild-type spores and unheated cC 1 spores revealed very few single-strand breaks (< 1 per 20 kb) in the DNA. In contrast, the DNA from heated a-0-spores had more than 10 single-strand breaks per 20 kb.These data suggest that binding of at/,3-type SASP to spore DNA in vivo greatly reduces DNA damage caused by heating, increasing spore heat resistance and long-term survival. While the precise nature of the initial DNA damage after heating of 13-spores that results in the single-strand breaks is not clear, a likely possibility is DNA depurination. A role for oc/13-type SASP in protecting DNA against depurination (and thus promoting spore survival) was further suggested by the demonstration that these proteins reduce the rate of DNA depurination in vitro at least 20-fold. Britain.tures to those at lower temperatures suggests that spores should be able to survive for years at common environmental temperatures (5, 24). Indeed, there are reports of spore survival over thousands of years (although obviously no details on spore population survival) (34). If spores are to survive for these extended times, their DNA must somehow be protected against the accumulation of potentially lethal damage during these periods, as the absence of metabolism and ATP in dormant spores precludes DNA repair (29,30,32). As noted above, spore DNA is protected against damage by UV radiation through the binding of a/1-type SASP. Other possible types of DNA damage are oxidative damage to bases and base loss through depurination; either of these types of DNA damage can lead to a mutagenic or lethal change in the DNA (7,10).DNA depurination appears to be of special significance, as this process is water catalyzed at neutral pH, with the rate of depurination rising dramaticall...

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“…Perhaps regulatory approvals for some non-medical applications of phage therapy (biocontrol), which have been granted in the United States, may be of interest. The Food and Drug Administration has amended the US food additive regulations to provide for the safe use of bacteriophages on ready-to-eat meat against Listeria monocytogenes [58]. The idea that ready-to-eat meat can be treated if contaminated with Listeria bacteria, while a doctor could not get a pharmaceutical grade phage therapy product when faced with a patient suffering listeriosis, strikes these authors as absurd, especially considering the recalls of various foods due to contamination with Listeria.…”

Section: Rediscovering Phage Therapymentioning

confidence: 99%

“…The patient's symptoms ceased after a single treatment, and he made a complete recovery. Dr. d'Herelle's anti-dysentery phage was then administered to three additional patients, all of whom began to recover within 24 hours of treatment [58]. In 1923, two physicians from Baylor University's College of Medicine reported successful results from one of their phage therapy trials conducted in United States, and concluded that "the bacteriophage holds enormous possibilities as a new weapon for fighting infectious disease" [31].…”

Section: Phage Historymentioning

confidence: 99%

Bacteriophage therapy: a potential solution for the antibiotic resistance crisis

Golkar

Bagasra

Pace

2014

J Infect Dev Ctries

492

337

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The emergence of multiple drug-resistant bacteria has prompted interest in alternatives to conventional antimicrobials. One of the possible replacement options for antibiotics is the use of bacteriophages as antimicrobial agents. Phage therapy is an important alternative to antibiotics in the current era of drug-resistant pathogens. Bacteriophages have played an important role in the expansion of molecular biology and have been used as antibacterial agents since 1966. In this review, we describe a brief history of bacteriophages and clinical studies on their use in bacterial disease prophylaxis and therapy. We discuss the advantages and disadvantages of bacteriophages as therapeutic agents in this regard.

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Mutation and killing of Escherichia coli expressing a cloned Bacillus subtilis gene whose product alters DNA conformation

Cited by 18 publications

References 35 publications

Lipids in the inner membrane of dormant spores of Bacillus species are largely immobile

Lipids in the inner membrane of dormant spores of Bacillus species are largely immobile

Prevention of DNA damage in spores and in vitro by small, acid-soluble proteins from Bacillus species

Bacteriophage therapy: a potential solution for the antibiotic resistance crisis

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