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J Mandelstam

Publications and source records attributed to J Mandelstam.

At least 37 records · Page 2Linked to original sources
PubMed1984

Use of temperature-sensitive mutants to study gene expression of two closely linked sporulation loci in Bacillus subtilis.

The spoOB and spoIVF loci are contiguous on the chromosome of Bacillus subtilis, so that genes in these loci may be parts of a single polycistronic operon. Temperature-sensitive strains having mutations in these loci were isolated, and temperature-shift experiments were carried out to investigate expression of the genes. The temperature-sensitive periods of spoOB mutants extended from the beginning of sporulation until the end of the stage II. The temperature-sensitive periods of spoIVF strains were during stage IV of sporulation. Therefore, although the spoOB and spoIVF loci are contiguous on the chromosome it is unlikely that genes in them are parts of a single polycistronic operon.

Bacillus subtilis↗
PubMed1984

Genetic and phenotypic characterization of a cluster of mutations in the spoVA locus of Bacillus subtilis.

Twenty-nine mutants blocked during stage V of sporulation have been isolated following directed mutagenesis of the lys-1 region of the Bacillus subtilis 168 chromosome. All of a sample of eight mutants tested are unaffected in sporulation marker events up to stage IV but did not produce dipicolinic acid. They produced stable 'phase white' spores that were released from the mother cell, and were partially resistant to toluene and lysozyme but sensitive to chloroform and heat. Mutation spoV A89, known to be in the lys-1 region, showed similar phenotypic characteristics. Three-factor transformation crosses and recombination indices showed that the new mutations and spoV A89 lie in a single linkage group, which maps between lys-1 and another sporulation locus, spoIIA. The size of the spoV A locus is such that it probably contains several genes, and these may be contiguous with the cluster of genes included within the spoIIA locus.

Bacillus subtilis↗
PubMed1984

Cloning of the Bacillus subtilis spoIIA and spoV A loci in phage phi 105DI:1t.

A 6.95 kb HindIII-generated DNA fragment from Bacillus subtilis 168 was inserted into the DNA of phage phi 105DI:1t. The recombinant phage (phi 105DS1) contained DNA of 33.8 kb as compared with 35.2 kb for phi 105DI:1t and 39.2 kb for the wild-type phage. In the presence of helper phage, phi 105DS1 complemented both spoIIA and spoV A mutations in B. subtilis.

Bacillus subtilis↗
PubMed1983

Variety of sporulation phenotypes resulting from mutations in a single regulatory locus, spoIIA, in Bacillus subtilis.

Closely linked mutations in either of the two putative genes of the sporulation locus spoIIA can affect, in quite diverse ways, spore incidence, the production of alkaline phosphatase and DNAase, and the stability of the cells in sporulation medium. It is concluded that the locus has a regulatory function affecting the activation or induction of at least two, and possibly more, sporulation-associated operons.

Alkaline Phosphatase↗
PubMed1983

Cloning of the Bacillus subtilis lys and spoIIIB genes in phage phi 105.

The lys gene of Bacillus subtilis was inserted into prophage phi 105. The recombinant phage (phi 105dlys) contained DNA which was about 2 MDal smaller than the wild-type phage DNA, and the phage particles had no tails. The phage did not plaque but, when provided with tails in vitro, it transduced both lys-1 and lys-3 strains of B. subtilis to Lys$. The lys$ gene was located on a 2.5 MDal EcoRI restriction fragment. Subsequently this phage was phi 105 105dspoIIIB, was also defective, i.e. without tails. The DNA was 1.5 MDal smaller than the wild-type phage DNA and the spoIIIB2$ gene was located on a 3 MDal EcoRI fragment. When provided with tails in vitro, phage phi 105dspoIIIB transduced cells of a spoIIIB2 recipient to Spo$. In these transductants the spoIIIB2 mutation was complemented, and the cells sporulated normally.

Bacillus subtilis↗
PubMed1981

Oxolinic acid-resistant mutants of Bacillus subtilis.

Two mutants of Bacillus subtilis have been independently selected for resistance to oxolinic acid, and inhibitor of DNA gyrase. The mutations, designated oxr-1 and oxr-2, map very close to one another but are clearly separated from mutations in the genes for DNA gyrase. Many of the phenotypic properties of the mutants differ from those of a strain containing the gyrA mutation described by other workers. In particular, the oxr strains are as sensitive as the wild-type to inhibition by nalidixic acid on solid medium. In addition, experiments with DNA synthesis in toluenized cells show that the enzyme of the gyrA mutant is resistant to oxolinic acid, whereas DNA synthesis in the oxr mutants is as sensitive as it is in wild-type preparations. It is concluded that resistance to oxolinic acid is not due to an alteration in the DNA gyrase, but is more probably the result of an impaired uptake of the inhibitor. Although growth of the mutants on agar plates is inhibited at high concentrations of oxolinic acid, lower concentrations (1-2 microgram ml-1) can be used to distinguish them from the wild-type. The oxr-1 and oxr-2 mutations define a new genetic locus and can be used as genetic markers in B. subtilis.

Bacillus subtilis↗
PubMed1981

Inhibition of sporulation by DNA gyrase inhibitors.

The effects of oxolinic acid and novobiocin - which respectively inhibit the A and B subunits of DNA gyrase, and therefore inhibit DNA synthesis - have examined in sporulating cultures of Bacillus subtilis. Although both inhibitors prevent sporulation, this is not due to inhibition of DNA synthesis. Instead, they affect protein synthesis generally, though weakly, but have very marked effects on the formation of individual enzymes. These effects are reproducible, but the type of enzyme that will be affected is not predictable. The results point to an involvement of DNA gyrase in the transcription of some genes. This is suggested as the reason for the effect of the inhibitors on spore formation, which they block mainly at Stage O-I.

Bacillus subtilis↗
PubMed1980

Germination properties as marker events characterizing later stages of Bacillus subtilis spore formation.

At various stages during spore formation sporangia were shocked by cold treatment or with toluene, and the germination requirements of the prespores were examined. Up to 5 h after induction of sporulation (t5) germination was spontaneous; i.e., it occurred without any added germinants. After t5, during stages V and VI, the capacity for spontaneous germination diminished progressively, and the spores acquired a need for externally added germinants. At t6 this need was satisfied by either L-alanine or a mixture of KCl, glucose, and fructose. By t8, the latter response had disappeared. The spores germinated only with L-alanine, and the response was much slower. Experiments with chloramphenicol showed that the germination properties of the spores appearing between t6 and t8 were the expression of events in protein synthesis that had occurred before t5. Although the germination requirements developed at about the same time as heat resistance, they could be dissociated from heat resistance in wild-type and mutant cells. The germination properties of the developing spores are additional marker events characterizing the later stages of sporulation, as follows: (i) spontaneous germination (up to the end of stage IV); (ii) germination requirements that are satisfied by KCl-glucose-fructose or L-alanine (stage V); and (iii) slow germination response with L-alanine only (stage VI).

Alanine↗
PubMed1980

Temporal dissociation of late events in Bacillus subtilis sporulation from expression of genes that determine them.

During sporulation in replacement medium, resistance to toluene to heating at 65 degrees C, to lysozyme, and to heating at 80 degrees C appeared in sequence between 4 and 8 h after the induction of sporulation (i.e., between t4 and t8). The addition of sufficient chloramphenicol at t4.5 to prevent protein synthesis nevertheless allowed the emergence of all of these types of resistance except lysozyme resistance. The numbers of spores with these types of resistance (lysozyme resistance again excepted) increased about fourfold when phenylmethylsulfonyl fluoride (an inhibitor of serine protease activity) was also present. Thus, the observed increases in resistance in the 2 h after the addition of chloramphenicol resulted from the utilization of preformed protein elements. Dipicolinate did not seem to be a determining factor in the development of any of these forms of resistance. Electron micrographs showed that inhibition of protein synthesis did not prevent deposition of the outer layers of the spores. Lysozyme resistance developed differently; synthesis of the relevant proteins began later (t5), and continued synthesis was necessary up to t8. Some processing of proteins made earlier was a prerequisite for lysozyme resistance. Therefore, it appears that from the viewpoint of regulation, the expression of the genes and the production of the proteins for resistance to toluene, heating at 65 degrees C, and heating at 80 degrees C are all stage IV sporulation events, although the resistance properties themselves appear only during stages V and VI. Lysozyme resistance is the only real late event among those examined. The germination characteristics of the spores, which are also late events, are discussed in this context, as they too are dependent on proteins that are synthesized much earlier.

Bacillus subtilis↗
PubMed1978

Extracellular manganese-stimulated deoxyribonuclease as a marker event in sporulation of Bacillus subtilis.

A considerable amount of Mn2+-stimulated DNAase (deoxyribonuclease) activity is released by Bacillus subtilis 168 during sporulation in a glucose-deficient medium; much smaller amounts are released during starvation for phosphate or nitrogen. Protein synthesis is required. Two forms of evidence are presented that production of the DNAase is associated with events late in stage II of sporulation. 19 Thymidine starvation, which inhibits the biochemical events associated with sporulation, also inhibits release of the DNAase. 2. Several asporogenous mutants blocked at stage II or earlier and unable to produce alkaline phosphatase (a stage-II event) do not produce the enzyme. Mutants blocked towards the end of stage II or later produce both enzymes. During sporulation of the wild-type strain, the DNAase appears about 1 h after alkaline phosphatase. The results suggest that production of the DNAase is controlled by a still-undiscovered stage-II genetic locus.

Bacillus subtilis↗
PubMed1976

Order of expression of genes affecting septum location during sporulation of Bacillus subtilis.

The mean volumes of stationary-phase cells of wild-type and asporogenous mutants of Bacillus subtilis have been measured. Mutants blocked at stage 0 of sporulation either produced cells that had the same volume as the developing sporangium or they divided to produce cells of one-half this volume. The order of expression of the genes affected by the mutations in these strains was determined by biochemical characterization and by construction of double sporulation mutants. Mutants that produced small cells were blocked at an earlier stage of sporulation than those that produced large cells. It is suggested that the following dependent sequence must occur before the formation of the prespore spetum: (i) the initiation of sporulation, (ii) a signal to block the final central division site, and (iii) a signal to activate a polar septum site.

Anti-Bacterial Agents↗
PubMed1976

Resporulation of outgrowing Bacillus subtilis spores.

Germinated spores of Bacillus subtilis were incubated in outgrowth medium and tested periodically for capacity to sporulate when suspended in sporulation medium. Concurrent measurements were made of deoxyribonucleic acid (DNA) content and numbers of cell division septa and nucleoids. Sporulation potential is shown to reach a peak at about 110 min at which time the chromosomes are probably well into the second round of replication. Experiments with nalidixic acid show that sporulation potential can be generated in the outgrowth medium even when DNA synthesis is largely prevented. Further experiments show that nalidixic acid apparently does not prevent the formation of DNA initiation complexes, which can subsequently function after resuspension in the sporulation medium. The results support those previously obtained with a temperature-sensitive DNA mutant which indicated that sporulation could only be induced at a specific stage of chromosome replication, and then only if the cells are in a state of nutritional "step-down".

Bacillus subtilis↗
PubMed1975

Production and possible function of serine protease during sporulation of Bacillus subtilis.

The production of extracellular protease during sporulation in Bacillus subtilis 168 was investigated. Two proteases are produced, an alkaline serine protease and a neutral metalloprotease. In vivo inhibition of the serine protease with phenylmethylsulfonylfluoride indicated that the metalloprotease was degraded by the serine protease during sporulation. The experiments with phenylmethylsulfonylfluoride also show that the serine protease is necessary for the sequential process of sporulation and that it is required continuously for the first 2 to 3 h of the 8-h process.

Alkaline Phosphatase↗
PubMed1975

Criteria for categorizing early biochemical events occurring during sporulation of Bacillus subtilis.

Two criteria are suggested for assessing the relevance of biochemical events occurring early in sporulation. The first is thymidine starvation, a condition known to inhibit sporulation. This also inhibits the production of metalloprotease, serine protease, and ribonuclease; alpha-amylase production, however, is unaffected. The second is the effect of a regulator mutation which increases the production of the proteases. In the mutant, ribonuclease is produced in correspondingly large quantities whereas alpha-amylase production is unaffected. We conclude that, whereas the serine protease is part of the main sequence of events leading to formation of the spore, the metalloprotease is a side effect, i.e., connected with the main sequence but not part of it. Ribonuclease could, on present evidence, be either in the main sequence or a side effect associated with it. Amylase, however, seems to be separately regulated and neither directly nor indirectly connected with the sporulation sequence.

Alkaline Phosphatase↗