Oxygen Toxicity and the Superoxide Dismutase

Oxygen caused an increase in the amount of superoxide dismutase in Escherichia coli B but not in Bacillus subtilis. E. coli B cells, induced by growth under 100% O(2), were much more resistant to the lethal effects of 20 atm of O(2) than were cells which contained the low uninduced level of this enzyme. In contrast, B. subtilis, which could not respond to O(2) by increasing its content of superoxide dismutase, remained equally sensitive to hyperbaric O(2) whether grown under 100% O(2) or areobically. The catalase in these organisms exhibited a reciprocal response to oxygen. Thus, the catalase of E. coli B was not induced by O(2), whereas that of B. subtilis was so induced. These results are consistent with the view that superoxide dismutase is an important component of the defenses of these organisms against the toxicity of oxygen, whereas their catalases are of secondary importance in this respect. The ability of streptonigrin to generate O(2) (-), by a cycle of reduction followed by spontaneous reoxidation, has been verified in vitro. It is further observed that E. coli B which contain the high induced level of superoxide dismutase were more resistant to the lethality of this antibiotic, in the presence of oxygen, than were E. coli B which contained the low uninduced level of this enzyme. This difference between induced and uninduced cells was eliminated by the removal of O(2). These results are consistent with the proposal that the enhanced lethality of streptonigrin under aerobic conditions may relate to its in vivo generation of O(2) (-) by a cycle of reduction and spontaneous reoxidation. In toto, these observations lend support to the hypothesis that O(2) (-) is an important agent of oxygen toxicity and that superoxide dismutase functions to blunt the threat posed by this reactive radical.     

Aspartokinase Activity and the Developmental Cycle of Myxococcus xanthus

The relationship between aspartokinase activity and fruiting body formation in Myxococcus xanthus was investigated. Two required amino acids, methionine and isoleucine, which stimulated the enzyme in vitro also inhibited fruiting body formation when added to 0.1% Casitone agar. Threonine, a potent feedback inhibitor of the aspartokinase, completely reversed the effects of methionine and isoleucine both on enzyme activity and fruiting body formation. A mutant, M. xanthus FB-S, which had the unusual property of forming fruiting bodies on 1.0% Casitone agar, also exhibited an altered regulation of aspartokinase activity. Spermidine, which is a strong stimulator of the enzyme in vitro, interfered with the developmental cycle of both M. xanthus FB and FS-S. During glycerol induction of myxospores the level of aspartokinase dropped more than 75% during the first hour. These data indicate a strong correlation between aspartokinase activity and the induction of the developmental cycle in M. xanthus. It is suggested that the decrease in aspartokinase activity results in diaminopimelic acid starvation, blockage of cell wall growth, and subsequent induction of the developmental cycle.     

Macromolecular Synthesis in Bacillus subtilis During Development of the Competent State

Rates of deoxyribonucleic acid, ribonucleic acid, and protein synthesis were examined in purified competent cells of Bacillus subtilis during the development of the transformable state. To become competent, a cell must depart from the normal course of vegetative growth and pass through a precompetent phase beginning as early as 90 to 180 min before the appearance of transformability. While in the precompetent state, the cell decreases its rate of deoxyribonucleic acid synthesis and lowers its ratio of ribonucleic acid synthesis to protein synthesis. This altered pattern of synthesis eventually leads to a decreased buoyant density of precompetent cells. Once a cell has become both precompetent and low in density, it can be converted to a competent (transformable) cell. The early alterations in macromolecular synthesis were found in two competence regimens, one utilizing a nutritional step-down and one free of such a shift. The data imply that the precompetent state is a generalized characteristic of the B. subtilis transformation system and is not specific to the procedure used to allow competence development. Since precompetence-specific events occur very early in a competence regimen, we conclude that the induction of precompetence is unrelated to sporulation or a nutritional shift.     

Heat-sensitive Step in Deoxyribonucleic Acidmediated Transformation of Bacillus subtilis

Over 90% of the competent cells in a population of Bacillus subtilis lost their competence after being heated to 50 C for 5 min. There was only a slight loss in the number of transformants if the culture was heated for 5 min after the termination of transformation, but 90% of the transformants were lost after 1 hr at 50 C. The population as a whole grew at a slightly faster rate at 50 C than at 32 C. We postulate that a heat-labile factor is required for the uptake or retention (or both) of deoxyribonucleic acid (DNA) in the cell, since uptake of ³²P-DNA into a deoxyribonuclease-resistant form was inversely proportional to the time of exposure to heat. Cells that had lost competence after being heated did not regain their competence for at least several hours, although other cells in the population became competent. These data suggest that the heat-labile factor required for competence is synthesized only once during the period that a cell remains competent.     

Purification and characterization of isocitrate lyase fromEscherichia coli

Isocitrate lyase has been purified to homogeneity, as determined by SDS-polyacrylamide gel electrophoresis and subsequent silver staining, fromEscherichia coli D₅H₃G₇. The enzyme was found to have a subunit molecular weight of 48,000 and a native molecular weight of 188,000 as determined by gel filtration chromatography. Thus, the enzyme appears to have tetrameric structure. The isoelectric point was determined to be 4.6, and the enzyme displayed a pH optimum at 7.3. The K_m of isocitrate lyase forthreo-Ds-isocitrate was determined to be 8 M. The purification procedure is highly reproducible and results in a 39% net yield of purified protein.     

Molecular Genetic Analysis of the DILUTE-SHORT EAR ( d-se) Region of the Mouse

Genes of the dilute-short ear (d-se) region of mouse chromosome 9 comprise an array of loci important to the normal development of the animal. Over 200 spontaneous, chemically induced and radiation-induced mutations at these loci have been identified, making it one of the most genetically well-characterized regions of the mouse. Molecular analysis of this region has recently become feasible by the identification of a dilute mutation that was induced by integration of an ecotropic murine leukemia virus genome. Several unique sequence cellular DNA probes flanking this provirus have now been identified and used to investigate the organization of wild-type chromosomes and chromosomes with radiation-induced d-se region mutations. As expected, several of these mutations are associated with deletions, and, in general, the molecular and genetic complementation maps of these mutants are concordant. Furthermore, a deletion breakpoint fusion fragment has been identified and has been used to orient the physical map of the d-se region with respect to the genetic complementation map. These experiments provide important initial steps for analyzing this developmentally important region at the molecular level, as well as for studying in detail how a diverse group of mutagens acts on the mammalian germline.     

Early development of johnny darter,Etheostoma nigrum,and fantail darter,E. flabellare,with a discussion of its ecological and evolutionary aspects

Synopsis Fantail darters,Etheostoma flabellare, have larger eggs with greater yolk volume than johnny darters,E. nigrum, and have a longer embryo period. This longer embryo period is associated with accelerated finfold differentiation, producing a larger, better formed individual at the onset of exogenous feeding. Developmental differences among these two species and the rainbow darter,Etheostoma caeruleum, primarily in timing of ontogenetic events, are described in detail. These differences in timing are considered largely early developmental phenomena as they affect embryos, larvae, and aievins but apparently not juveniles or adults. Demographic aspects of early life history were assessed using data from studies of Ontario streams. Differences in early survival among species appear to play an important role in recruitment of mature females, but cannot be attributed to parental care or egg size differences.     

Active entry of stream benthic macroinvertebrates into the water column

Field experiments investigated the possible active entry of stream benthos into the water column. Over a 1-year period, sediment baskets were suspended for 24 hours in a stream pool so that only swimming or floating organisms, essentially unaided by current, could colonize them. A variety of benthos, including taxa characteristic of riffles, colonized the baskets, with colonization highest in late summer and negligible in winter.A modified drift net towed through the pool was used to quantitatively sample benthic animals actually in the water column. Nighttime tows captured a diverse, abundant fauna and indicated densities substantially higher than invertebrate drift densities reported in the literature. Daytime tows yielded little. Estimated percentages of the benthos in the water column at a given time were generally < 1.0%.These findings suggest that not all invertebrate drift is the result of passive mechanical removal from the substrate by current.