Cloning and expression of Candida albicans ADE2 and proteinase genes on a replicative plasmid in C. albicans and in Saccharomyces cerevisiae.

Summary A plasmid vector (denoted pRC2312) was constructed, which replicates autonomously in Escherichia coli, Saccharomyces cerevisiae and Candida albicans. It contains LEU2, URA3 and an autonomously replicating sequence (ARS) from C. albicans for selection and replication in yeasts, and bla (ampicillin resistance) and ori for selection and replication in E. coli. S. cerevisiae AH22 (Leu^–) was transformed by pRC2312 to Leu^– at a frequency of 1.41 × 10⁵ colonies per g DNA. Transformation of C. albicans SGY-243 (Ura-) to Ura⁺ with pRC2312 resulted in smaller transformant colonies at a frequency of 5.42 × 10³ per g DNA where the plasmid replicated autonomously in transformed cells, and larger transformant colonies at a frequency of 32 per g DNA, in which plasmid integrated into the genome. Plasmid copy number in yeasts was determined by a DNA hybridization method and was estimated to be 15±3 per haploid genome in S. cerevisiae and 2–3 per genome in C. albicans replicative transformants. Multiple tandem integration occurred in integrative transformants and copy number of the integrated sequence was estimated to be 7–12 per diploid genome. The C. albicans ADE2 gene was ligated into plasmid pRC2312 and the construct transformed Ade^– strains of both C. albicans and S. cerevisiae to Ade⁺. The vector pRC2312 was also used to clone a fragment of C. albicans genomic DNA containing an aspartic proteinase gene. C. albicans transformants harboring this plasmid showed a two-fold increase in aspartic proteinase activity. However S. cerevisiae transformants showed no such increase in proteinase activity, suggesting the gene was not expressed in S. cerevisiae.     

Receptor-independent catabolism of low density lipoprotein. Involvement of the reticuloendothelial system

In this study we examine the effects of intravenous ethyl oleate emulsions on the metabolism of native and cyclohexanedione-modified human low density lipoprotein in rabbits. Treatment produced a highly significant fall in receptor-independent catabolism as measured by the fractional clearance rate of cyclohexanedione-modified low density lipoprotein. Receptor-dependent catabolism (the difference between the fractional clearance rates of native and cyclohexanedione-modified low density lipoprotein) was variably affected with some animals showing a decrease in receptor activity. These data suggest that the reticuloendothelial system makes a substantial contribution to receptor-independent low density lipoprotein catabolism in the rabbit.     

Comparison of the chromosomes of Triticum timopheevi with related wheats using the techniques of C-banding and in situ hybridization

Summary The chromosomes of the tetraploid wheats Triticum timopheevi (Genome AAGG) and T. araraticum (Genome AAGG) were C-banded at mitosis. The identity of the banded and unbanded chromosomes was then established by firstly making comparisons with the hexaploid species T. zhukovskyi which has the genome formula AAAAGG. Secondly, the meiotic pairing in F₁ hybrids between T. timopheevi and diploid wheats was examined by means of C-banding. The results showed that the banded chromosomes belonged to the G genome, while the unbanded chromosomes belonged to the A genome. Only one of the two pairs of satellited chromosomes had strong heterochromatic bands. The relationship between the genomes of T. timopheevi and T. dicoccum (Genome AABB) was then assessed at meiosis in hybrids between these species, using the techniques of C-banding and in situ hybridisation of a cloned ribosomal RNA gene probe. It was concluded that there were differences both in the amount and distribution of heterochromatin and also translocation differences between the species.     

Dimeric epoxy fatty acid methyl esters: formation,chromatography and mass spectrometry

The formation of dimers is reported from the thermal treatment of a series of epoxy fatty acid methyl esters. These compounds were isolated from the reaction mixture by steric exclusion chromatography and were subsequently characterised by their high resolution electron impact and ammonia chemical ionisation mass spectra. The spectra were consistent in each case with the presence of a mixture of four possible positional isomers each containing an ether bridge linking a pair of fatty acid methyl esters across the carbon chains, with a keto group on a carbon adjacent to the bridge on one of the esters.     

Genetic Instability in DROSOPHILA MELANOGASTER: The Induction of Specific Chromosome 2 Deletions by MR Elements

We describe the spontaneous induction of deletions by MR elements at the l(2)gl, net, pr and cn loci. The frequent induction of l(2)gl deletions mimics the high frequency of l(2)gl alleles found in wild populations of D. melanogaster. We suggest that these and other data that we present militate for the conclusion that, in the wild, autonomous MR elements occur and function as mutators. We contend that MR elements are not simply the by-products of hybridization between wild and laboratory strains.     

Zwitterionic dipoles as a dielectric probe for investigating head group mobility in phospholipid membranes

For phospholipid membranes with zwitterionic head groups, the dipole can be considered as a specific label for tracing the changes in the dynamic behaviour of this region of the bilayer in its various phases. Measurements of the dielectric properties of fully hydrated 1,2-dipalmitoyl-sn-glycero-3-phosphocholine bilayers in the frequency range 1--50 MHz show a dispersion which is attributed to the motion of the phosphocholine dipoles in the plane of the bilayers. When the temperature is varied, both the permittivity and loss factor increase sharply at the pretransition (35 degrees C) and the main transition (42 degress C). The relaxation time and amplitude were also determined for this dispersion and these further reflect the structural changes occurring with temperature. The relaxation times varied between 4 ns at 30 degrees C and 2.3 ns at 50 degrees C. Due to steric hindrances a restriction in the angle of head group rotation occurs at lower temperatures but is greatly reduced above the main transition.     

Acetylcholine--inhibition of transmitter release from adrenergic nerve terminals mediated by muscarinic receptors.

The evidence is reviewed for the presence of muscarinic receptors on the sympathetic nerves to blood vessels. Activation of these receptors by acetylcholine in doses that are too small to affect the smooth muscle cells directly inhibits the release of norepinephrine evoked by electric impulses or potassium ions. This inhibitory action of acetylcholine is prevented by muscarinic blocking agents and is probably due to hyperpolarization of the adrenergic nerve terminals.     

Metabolic modulation of neurotransmitter release--adenosine, adenine nucleotides, potassium, hyperosmolarity, and hydrogen ion.

Evidence has accumulated that several factors, which have been proposed as mediators of exercise hyperemia, can modulate adrenergic neurotransmission in blood vessels. Adenosine and the adenine nucleotides depress the response of isolated blood vessels of the dog to nerve stimulation more than that to exogenous norepinephrine; this difference is explained by a decreased release of the neurotransmitter. Potassium, hyperosmolarity, and acidosis also depress adrenergic neurotransmission in isolated veins. These results are consistent with the hypothesis that metabolic changes in the vicinity of the adrenergic neuroeffector junction are capable of decreasing the output of neurotransmitter to the blood vessels in the exercising muscle.     

Mannitol production in fungi during glucose catabolism.

The levels of phosphofructokinase (EC 2.7.1.11) and mannitol-1-phosphate dehydrogenase (EC 1.1.1.17) have been determined in a number of Mucor and Penicillium species. Mannitol-1-phosphate dehydrogenase was found in only one species of mucor, Mucor rouxii, and this with a specific activity much lower than that found in Penicillium species. All of the fungi tested in the Ascomycetes class exhibited mannitol-1-phosphate dehydrogenase activity. Interference from both mannitol-1-phosphate dehydrogenase and NADH oxidase (EC 1.6.99.5) caused some difficulty initially in detecting phosphofructokinase in Penicillium species; the Penicillium phosphofructokinase is very unstable. Penicillium notatum accumulates mannitol intracellularly; detection of mannitol-1-phosphate dehydrogenase and mannitol-1-phosphatase (EC 3.1.3.22) activity in cell-free extracts indicates that the mannitol is formed from glucose via fructose-6-phosphate and mannitol-1-phosphate; no direct reduction of fructose to mannitol could be detected. The mannitol-1-phosphate dehydrogenase was specific for mannitol-1-phosphate and fructose-6-phosphate; NADP+(H) could not replace NAD+(H). The phosphatase (EC3.1.3.22) exhibited a distinct preference for mannitol-1-phosphate as substrate; all other substrates tested exhibited less than 25% of the activity observed with mannitol-1-phosphate.