Polyamines and the Accumulation of Ribonucleic Acid in Some Polyauxotrophic Strains of Escherichia coli

The cellular accumulations of polyamines and ribonucleic acid (RNA) were compared in the polyauxotrophic mutants of Escherichia coli strain 15 TAU and E. coli K-12 RC(re1) met(-) leu(-). Putrescine, spermidine, and their monoacetyl derivatives were the main polyamines in both strains, when grown in glucose-mineral medium. No significant degradation of either (14)C-putrescine or (14)C-spermidine was found in growing cultures of strain 15 TAU, which requires thymine, arginine, and uracil for growth. Experiments with this organism showed that in a variety of different incubation conditions, which included normal growth, amino acid starvation, inhibition by chloramphenicol or streptomycin, or thymine deprivation, a close correlation was seen between the intracellular accumulation of unconjugated spermidine and RNA. In the presence of arginine, the antibiotics stimulated the production of putrescine and spermidine per unit of bacterial mass. Deprivation of arginine also resulted in an increase in the production of putrescine per unit of bacterial mass, most of which was excreted into the growth medium. However, in this system the antibiotics reduced the synthesis of putrescine. Furthermore, streptomycin caused a rapid loss of cellular putrescine into the medium. The latter effect was not seen in anaerobic conditions or in a streptomycin-resistant mutant of 15 TAU. Methionine added to the growth medium of growing TAU not only markedly increased the total production of spermidine, but also increased both the intracellular concentration of spermidine and the accumulation of RNA. Exogenous spermidine extensively relaxed RNA synthesis in amino acid-starved cultures of 15 TAU. Analysis in sucrose density gradients showed that the RNA accumulated in the presence of spermidine was ribosomal RNA.Cells of E. coli K-12 RC(rel) met(-) leu(-), grown in a complete medium, had approximately the same ratio of free spermidine to RNA as did strain 15 TAU. However, the relaxed strain showed a much lower ratio of putrescine to spermidine than the stringent 15 TAU. Omission of methionine stopped spermidine synthesis and markedly increased both the intracellular accumulation and the total production of putrescine. It seems that a high intracellular level of spermidine acts as a feedback inhibitor in the biosynthesis of putrescine in this strain. The hypothesis that the intracellular concentration of polyamines may participate in the control of the synthesis of ribosomal RNA in bacteria is discussed.     

Intraspecific hybridization and its bearing on chromosome evolution inVicia narbonensis (Fabaceae)

Nine accessions ofVicia narbonensis, considered to be the wild progenitor of faba bean (Vicia faba), were investigated to ascertain the nature and extent of intraspecific karyotypic polymorphism. The chromosome complements resolved into four distinct types (A, B, C, D), and the meiotic data of F₁ hybrids (A × B, B × C, A × C) revealed that alteration in chromosome morphology is the result of segmental interchanges. The interchange complexes indicate that the parents differ from each other by 1 to 2 interchanges. It is also evident that karyotype B, and not A as previously reported, is the normal karyotype of the species, and A and C are single homozygotes for unequal interchange. The comparative karyomorphology of the parents and the hybrids, and of two interchange heterozygotes of four chromosomes each in F₁ hybrids of A × C shows that the chromosomes involved in the single interchange homozygotes (A, C) are not common and the breaks in both interchanges occurred in short and long arms of the involved chromosomes. Identification of the interchanged chromosomes in the complements and the frequency of ring and chain quadrivalents in the heterozygotes enabled location of the breakpoints. The present results provide probably the first example indicating that interchange homozygosity (A) is not only firmly established but also has enabled the species to spread further by adapting to a wide range of habitats. — The genetic relationships between A and D are very different. All seven chromosome pairs in D could be distinguished from A, and for that matter, B and C as well. From the meiotic pairing properties it is also amply clear that genome D is well differentiated from A and possibly B, and C, and deserves special status.     

Transfer of Brassica tournefartii (TT) genes to allotetraploid oilseed Brassica species (B. juncea AABB, B. napus AACC, B. carinata BBCC): homoeologous pairing is more pronounced in the three-genome hybrids (TACC, TBAA, TCAA, TCBB) as compared to allodiploids (TA, TB, TC)

For the transfer of genes from B. tournefortii (TT) to the allotetraploid oilseed brassicas, B. juncea AABB, B. carinata BBCC and B. napus AACC, B. tournefortii was first crossed with the three basic diploid species, B. campestris (AA), B. nigra (BE) and B. oleracea (CC), to produce the allodiploids TA, TB and TC. These were tetraploidized by colchicine treatment to produce the allotetraploids TTAA, TTBB and TTCC, which were further crossed with B. juncea and B. napus to produce three-genome hybrids with substitution-type genomic configurations: TACC, TBAA and TCAA. These hybrids along with another hybrid TCBB produced earlier, the three allodiploids, their allotetraploids and the four diploid parent species were studied for their male meiotic behaviour. The diploid parent and the allotetraploids (TTAA, TTBB and TTCC) showed regular meiosis although the pollen viability was generally low in the allotetraploids. In the allodiploids (TA, TB and TC) only some end-to-end associations were observed without any clearly discernible chiasmata or exchange points. Chromosomes involved in end-to-end associations were randomly distributed at the metaphase/anaphase-I stages. In contrast, the three-genome hybrids (TACC, TBAA, TCAA and TCBB) showed normal bivalents whose number exceeded the expected bivalent values. Bivalents arising out of homoeologous pairing were indistinguishable from normal pairs by their disjunction pattern but could be distinguished on the basis of the heteromorphy of the homoeologous chromosomes. The three-genome hybrids could be backcrossed to allotetraploid oilseed brassicas as they had some fertility. In contrast, the allodiploids could neither be selfed nor back-crossed. On the basis of their meiotic stability, in terms of more pronounced homoeologous pairing and fertility for backcrossing, the three-genome configurations provide the best possible situation for the introgression of alien genes from the secondary gene pool to the allotetraploid oilseed crops B. juncea, B. napus and B. carinata.     

Mating-induced loss of sex pheromone and sexual receptivity in insects with emphasis on Helicoverpa zea and Lymantria dispar

Mating in most species of insects leads to a transient or permanent loss in sexual receptivity of the females. Among moths, this loss of receptivity is often accompanied with a loss of the sex pheromone in the absence of calling, which also could be temporary or permanent. Most of the earlier work on changes in reproductive behavior after mating was done with Diptera in which sperm and/or male accessory gland secretions were shown to be responsible for termination of receptivity. In the corn earworm moth, Helicoverpa zea, mated females become depleted of pheromone and become nonreceptive to further mating attempts, but only for the remainder of the night of mating. A pheromonostatic peptide isolated from the accessory glands of males may be responsible for the depletion of pheromone, while the termination of receptivity is independently controlled. In the gypsy moth, Lymantria dispar, the changes in behavior following mating are permanent. In this species, the switch from virgin to mated behavior involves three steps: a physical stimulation associated with mating, transfer of viable sperm to the spermatheca, and commencement of oviposition. Signals generated by these factors operate through neural pathways and, unlike in H. zea, accessory gland factors seem not to be involved. © 1994 Wiley-Liss, Inc.

1 This article is a US Government work and, as such, is in the public domain in the United States of America.

     

Identification and characterization of a new disulfide isomerase-like protein (DsbD) in Escherichia coli.

Previous studies have established that DsbA and DsbC, periplasmic proteins of Escherichia coli, are two key players involved in disulfide bond formation. A search for extragenic mutations able to compensate for the lack of dsbA function in vivo led us to the identification of a new gene, designated dsbD. Lack of DsbD protein leads to some, but not all, of the phenotypic defects observed with other dsb mutations, such as hypersensitivity to dithiothreitol and to benzylpenicillin. In addition, unlike the rest of the dsb genes, dsbD is essential for bacterial growth at temperatures above 42 degrees C. Cloning of the wild-type gene and sequencing and overexpression of the protein show that dsbD is part of an operon and encodes an inner membrane protein. A 138 amino acid subdomain of the protein was purified and shown to possess an oxido-reductase activity in vitro. Expressing this subdomain in the periplasmic space helped restore the phenotypic defects associated with a dsbD null mutation. Interestingly, this domain shares 45% identity with the portion of the eukaryotic protein disulfide isomerase carrying the active site. We further show that in dsbD mutant bacteria the dithiol active sites of DsbA and DsbC proteins are mostly oxidized, as compared with wild-type bacteria. Our results argue that DsbD generates a reducing source in the periplasm, which is required for maintaining proper redox conditions. The finding that overexpression of DsbD leads to a Dsb- phenotype, very similar to that exhibited by dsbA null mutants, is in good agreement with such a model.     

Effect of long term feeding of rice bran oil upon lipids and lipoproteins in rats

The effects of feeding two levels of rice bran oil (RBO) on the growth, lipid parameters, and fatty acid composition of the plasma and liver of rats (Wistar strain) were compared with those produced on animals which had been fed the same levels of peanut oil (PNO). The control animals were fed synthetic diets containing 5 and 20% peanut oil (PNO) and the experimental groups were fed similar diets, containing the same level of rice bran oil (RBO). There was no significant difference with respect to the organ weights between the control and the experimental groups. In general, groups fed 20% oil gained more weight than groups fed 5% oil. The animals which received rice bran oil in their diet had, in general, comparatively lower levels of cholesterol, triglycerides and phospholipids. On the other hand, animals receiving 20% rice bran oil in their diet, showed an increase of 20% in high density lipoproteins (HDL-C), within 18 weeks (p<0.05), when compared to the animals fed with peanut oil. Similarly, low density lipoprotein cholesterol (LDL-C) and very low density lipoprotein cholesterol (VLDL-C) were lower in RBO-fed groups, than in the PNO-fed groups. There was, however, no significant differences in the cholesterol/phospholipid (C/P) ratio of the two groups. Analysis of plasma and of liver fatty acids indicated, in a general way, the type of fat consumed. There were no significant difference in the P/S ratio, nor any in the oleic/linoleic, oleic/stearic, palmitoleic/palmitic, oleic/palmitic, and oleic/palmitoleic ratios. Furthermore, levels of saturated (SAFA), monounsaturated (MUFA), and polyunsaturated (PUFA) fatty acids were identical in both the groups. Thus, our results suggest that feeding a high level of rice bran oil (RBO) has no deleterious effect on the growth and blood lipid profile of rats.Abbreviations PNO peanut oil - RBO rice bran oil - HDL-C high density lipoprotein cholesterol - LDL-C low density lipoprotein cholesterol - VLDL-C very low density lipoprotein cholesterol - SAFA saturated fatty acids - MUFA mono-unsaturated fatty acids     

Field-scale biofiltration of gasoline vapors extracted from beneath a leaking underground storage tank

Approximately 15000 L of unleaded gasoline werereleased into the surrounding vadose zone from aleaking underground storage tank. Initialremediation was by soil vapor extraction andcombustion which soon became cost prohibitive, asadded propane was required to reach the combustionlimit of the extracted vapors. As a cost effectivealternative, a field-scale compost based biofilterwas used in conjunction with soil vapor extractionto remediate the vadose zone. The biofilter wasconstructed on site using 4:1 diatomaceousearth:composted horse manure. Results of a fivemonth study showed that the biofilter removedapproximately 90% of total petroleum hydrocarbons(TPH) and >90% of the BTEX compounds (benzene,toluene, ethylbenzene, xylene), achieving thestringent permit requirements set at either 90% TPHreduction or less than 1.36 kg per day of volatileorganic compounds (VOC's) released to theatmosphere. The biofilter showed the capacity toreadily adapt to changing environmental conditionssuch as increased contaminant loading, andvariations in temperature and moisture. Thebacterial population in the biofilter was uniformlydiverse throughout the biofilter, suggesting that aconsortium of bacteria was needed for efficientbiodegradation. The cost of biofilter set up andoperation saved 90% in the first year alone of theoperating expenses incurred by soil vapor extractionand combustion.