Rapid Method for Measuring Extracellular Water in Yeast Preparations

A rapid procedure for the quantitative determination of extracellular water in bulk bakers' yeast was developed on the basis of the solute dilution principle. A reagent is prepared by synthesizing the diazonium ion of p-aminobenzoic acid and coupling it to peptone. This "azopeptone reagent" permits direct colorimetric measurement, which accounts for the rapidity and simplicity of the test. Potential errors due to osmotic effects are avoided by supplementing the reagent with saline and, more importantly, minimizing the duration of contact between reagent and cells. The new method has acceptable accuracy and precision, and may also be suitable for use with other microorganisms.     

In vitro adsorption of a hydrophobic mutagen to gastrointestinal mucus glycoprotein (mucin) and dietary fibre.

The adsorption of mutagens by some dietary fibres has been suggested as one mechanism by which dietary fibres protect against colorectal cancer. It is thought that these dietary fibres carry the mutagen out of the digestive tract, decreasing the effective mutagen concentration to which epithelial cells are exposed. The ability of gastrointestinal mucin to alter the extent to which the hydrophobic mutagen 1,8-dinitropyrene (DNP) adsorbs in vitro onto the insoluble dietary fibre alpha-cellulose, was investigated. It was found that crude and purified human ileal mucins themselves adsorbed DNP and decreased the adsorption of DNP onto alpha-cellulose. Purified mucin which had been treated with trypsin also adsorbed DNP. These studies suggest that in the digestive tract there would be competition for the adsorption of DNP between mucin and insoluble dietary fibres, such as alpha-cellulose. This factor must be considered in predictions about the distribution of hydrophobic, mutagenic carcinogens in the digestive tract and their role in the etiology of colorectal cancer.     

A mutant CHO-K1 strain with resistance to Pseudomonas exotoxin A and alphaviruses fails to cleave Sindbis virus glycoprotein PE2.

RPE.40, a mutant CHO-K1 strain selected for resistance to Pseudomonas exotoxin A, is defective in the production of infectious alphaviruses, although viruses are taken in and processed normally (J. M. Moehring and T. J. Moehring, Infect. Immun. 41:998-1009, 1983). To determine the cause of this defect, the synthesis of Sindbis virus proteins was examined. RPE.40 cells produced and glycosylated structural glycoprotein precursors PE2 and immature E1 normally. Mature E1 was formed, but PE2 was not cleaved to E2 and E3. PE2 instead was modified to a higher-molecular-weight form (PE2') in which the high-mannose oligosaccharides were processed to the complex form without proteolytic cleavage. The data suggest that the cleavage which produces E2 occurs within the trans-Golgi or in post-Golgi elements and is closely associated with the addition of sialic acid residues to the asparagine-linked oligosaccharides. RPE.40 cells make and release noninfectious Sindbis virions that contain PE2' and no detectable E2. These virions can be converted to an infectious form by treatment with trypsin. A defect in an intracellular endopeptidase activity in RPE.40 cells is postulated. Comparison of two Sindbis virus strains showed that the requirement for E2 in the virion to ensure infectivity is strain specific.     

Seasonal movement and spatial distribution of the sheep ked (Diptera: Hippoboscidae) on Wyoming lambs.

When populations of adult sheep ked, Melophagus ovinus (L.), infesting unshorn lambs were monitored at the University of Wyoming Paradise Farm during 1986, we determined the body regions on which keds would be found at various times of the year and their seasonal population trends for optimal sampling. Results suggested that ked populations were consistently greater on the ribs than on any other area of the lamb. No significant differences were detected for ked populations between sides of a lamb. Distinct and similar ked population trends over time occurred only in the rib, thigh, shoulder, hind leg, belly, and hind flank areas of the lambs, suggesting that a significant seasonal migration did not occur. Analyses for seasonal population fluctuations indicated that ked populations increased in the winter and spring, decreased in summer and then increased again in the fall. Thus, sampling for keds in the rib area at shearing, which begins in March in Wyoming and runs through mid-April, would be an opportune time to detect keds. At other times of the year, the rib area should be inspected for presence of sheep ked.     

The androgen receptor gene is located on a highly conserved region of the X chromosomes of marsupial and monotreme as well as eutherian mammals

The androgen receptor gene (AR), which is located on the long arm of the human X chromosome, was mapped by somatic cell analysis and in situ hybridization in marsupial and monotreme species. Both methods demonstrated that it was located on the X chromosome in each marsupial species, and also in the platypus. We conclude that this gene is part of a highly conserved region of the mammalian X, represented by the human Xq, which formed part of the X chromosome in a mammalian ancestor 150 million years ago. Since this gene is located proximally on the long arm of the monotreme X, which is G-band homologous to the Y and apparently exempt from X chromosome inactivation, the conservation of this region has evidently not depended on its isolation by X-Y differentiation or on X inactivation.     

Cerebellar [H]kainate and [H]ampa binding in dogs with congenital portosystemic encephalopathy

Previous studies have indicated that kainate and AMPA receptors are altered in cerebral cortex of dogs with chronic hepatic encephalopathy (HE). To ascertain whether receptors in dog cerebellum are similarly altered in HE [³H]kainate and [³H]-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) binding assays were performed on crude synaptosomal membranes prepared from cerebellar tissue from dogs with congenital portosystemic encephalopathy (PSE) and control dogs. There was no pathophysiologically relevant difference in the affinity or density of kainate or AMPA binding sites in PSE cerebellar tissue compared with control dogs. The failure to demonstrate alterations in these binding parameters in cerebellar tissue was expected as clinical signs of HE reflect cortical rather than cerebellar dysfunction.     

Activation of adenylate cyclase during swelling of S49 cells in hypotonic medium is not involved in subsequent volume regulation

Experiments in S49 mouse lymphoma cells indicate that adenylate cyclase activity is increased following swelling in hypotonic medium through a mechanism independent of the G-proteins which are involved in hormonal regulation of the enzyme. An intact actin cytoskeleton is apparently required for stimulation of adenylate cyclase by mechanical forces. It was hypothesized that this increase in cAMP may be involved in triggering subsequent volume regulatory events. Manipulation of intracellular CAMP content and protein kinase A activity in S49 cells prior to swelling or during the regulatory volume decrease following swelling provided no evidence of a significant role for CAMP in regulating the extent of initial volume increase or the subsequent regulatory volume decrease. Treatment of S49 cells with 10-200 µM miconazole, previously shown to inhibit adenylate cyclase activity, attenuated the initial volume increase with medium dilution and accelerated the rate of regulatory decrease in a dose-dependent and time-dependent manner. However, incubation with 100 µM miconazole for 20 min, which completely inhibited swelling-induced increases in cAMP content, had no significant effect on either the initial volume expansion or the extent of regulatory volume decrease.