Occurrence, isolation, and characterization of polyribosomes in yeast

This report details the procedural requirements for preparing cell-free extracts of yeast rich in polyribosomes. This enabled us to demonstrate the occurrence of polyribosomes in yeast, to show their role in protein synthesis, and to devise methods for their resolution and isolation. When certain precautions are met (the use of log phase cells, rapidly halting cell growth, gentle methods of disruption, sedimentation through exponential density gradients, etc.), individual polyribosome size classes ranging up to the heptosome can be fractionated and separated from their nearest neighbors. Larger size classes are resolved partially among themselves, free of smaller polyribosomes. This was confirmed by extensive electron micrographic studies of material from the various fractions obtained upon density gradient centrifugation of yeast extracts. Modifications of the gradients and procedure should allow fractionation and isolation of the larger polyribosomes, including those containing polycistronic messages. Yeast polyribosomes are disaggregated to single ribosomes by longer term grinding, cell disruption by the French pressure cell, the Hughes press, or by incubation with dilute RNAse. Yeast polyribosomes are active in the incorporation of amino acids into polypeptide; the single ribosomes exhibit only slight activity. The latter activity is probably due to the presence of a small fraction of monosomes still containing mRNA. Poly-U stimulates amino acid incorporation only in the single ribosomes.     

Genetic Analysis of Glutamate Transport and Glutamate Decarboxylase in Escherichia coli

The location of the Escherichia coli K-12 genes determining or regulating glutamate transport, and the location of the gene determining glutamate decarboxylase synthesis, were established by conjugation. The ability to grow on glutamate as the sole source of carbon and energy was used to select for glutamate transport recombinants. Two genes determining the ability to grow on glutamate as the sole source of carbon and energy were mapped. One (gltC) is located near mtl (mannitol), and the other (gltH) appears to be located between the gal (galactose) and trp (tryptophan) loci. The glutamate decarboxylase gene (gad) is strongly linked to gltC. The gltC(+) recombinants grow on glutamate much faster and accumulate this amino acid to a greater extent than do the gltH(+) recombinants. The gltH(+) gene functioned only in one female strain (P678), whereas the gltC gene functioned in all the female strains tested (P678, C600, W1).     

Eph Family Receptors and Their Ligands Distribute in Opposing Gradients in the Developing Mouse Retina

The Eph family of receptor tyrosine kinases and their ligands can be divided into two specificity subclasses: the Eck-related receptors and their GPI-anchored ligands, and the Elk-related receptors and their transmembrane ligands. Previous reports demonstrated that Eck- and Elk-related receptors in the retina distribute in high temporal–low nasal and high ventral–low dorsal gradients, respectively. While others have focused on complementary ligand gradients in the retinal axon target, the tectum, we report that ligands from each subclass also distribute in gradients opposing those of their corresponding receptors within the retina itself. Moreover, ligand gradients in the retina precede ganglion cell genesis. These results support an intraretinal role for Eph family members in addition to their previously proposed role in the development of retinotectal topography. The distinct distributions of Eph family members suggest that each subclass specifies positional information along independent retinal axes.     

Extremely short 20-33 nucleotide introns are the standard length in Paramecium tetraurelia.

Paramecium tetraurelia has the shortest known introns as its standard intron length. Sequenced introns vary between 20 and 33 nucleotides in length. The intron sequences were discovered in genomic sequences coding for a variety of different proteins, including phosphatases, kinases, and low-molecular weight GTP-binding proteins. All intron sequences begin with the conserved dinucleotide GT and end with the conserved dinucleotide AG. The sequences are more AT rich than the Paramecium coding sequences. The identified sequences were confirmed as introns by sequencing several cDNA fragments. We report here analysis of the characteristics of 50 separate introns, including size, base composition, and a consensus sequence.     

Use of recombinant interferon-α in Human Immunodeficiency Virus (HIV)-infected individuals

Rationale and objective Interferon alpha (IFN-) has anti-retroviral activity and is a possible HIV infection-limiting factor. The aim of this work is to prevent or delay disease progression in asymptomatic Human Immunodeficiency Virus (HIV) carriers.Design and interventions Recombinant IFN alpha-2b (3×10⁶ IU 3 times weekly) was compared. to no treatment (control) in a randomized trial. Endpoints were: (i) appearance of any CDC group IV symptoms and (ii) disease progression (which excluded shifts to group IVC2 or reversible IVA, or IVB). The trial lasted from October 1987 to February 1992.Setting The trial was performed at the Santiago de las Vegas sanatorium, a specialized institution for the care of HIV-infected and AIDS patients.Population Subjects were anti-HIV-1 seropositive, Western blot-confirmed, asymptomatic (CDC group II), or with generalized lymphadenopathies (CDC group III). The groups had 79 (control) and 71 (IFN) patients.Main results Long-term IFN- treatments significantly reduced the proportion of patients who shifted to any group IV (control: 46/79; IFN: 14/71;p<0.001) or developed AIDS (control: 27/79; IFN: 12/71;p<0.05). IFN also delayed progression to AIDS (95% confidence interval for 0.5 probability of progression) from 67–83 to 116–180 months after infection. The IFN group had significantly less opportunistic infections and non-infectious complications. CD4 cell count and hemoglobin decreased in the control but not in the IFN group. Fewer IFN-treated patients developed positive serum HIV antigen detection.Conclusion IFN alpha treatment during the early stages of infection seems to be beneficial to the patients.Abbreviations CI confidence interval - AIDS Acquired Immunodeficiency syndrome - HIV Human Immunodeficiency Virus - IFN Interferon - CDC Center for Disease Control (USA) - SD standard deviation     

Assessment of lead availability in soils contaminated by mine spoil

The uptake of lead by two contrasting plant species, radish and red fescue, grown in soils contaminated by mine spoil was investigated. Uptake was found to be poorly correlated either with pH or total Pb concentration in the soils. By contrast, a good correlation was obtained, particularly for red fescue, between Pb uptake and Pb concentration in the solution of equilibrated soil suspensions over a wide range of soil pH, total soil Pb and soil solution Pb concentration.Calculations suggested a similar order of magnitude in the total amounts of Pb taken up by the plants and Pb in the soil solution of the root zone, justifying the latter as a good index of Pb-availability. ei]Section editor: A C Borstlap     

On the probability of loss of new mutations in the presence of linkage disequilibrium

A new selectively neutral mutation occurs in a multilocus genetic background that has achieved a stable equilibrium at which there is a linkage disequilibrium. Perturbation techniques are applied to an extension of the branching process formulation of Fisher in order to address the question of extinction probabilities. We show that under appropriate conditions the probability of extinction of the new mutant is increased by the existence of linkage disequilibrium in the genetic background.Research supported in part by NIH grant GM 28016     

Root Morphology and Nitrogen Uptake of Maize Simultaneously Supplied with Ammonium and Nitrate in a Split-root System

We studied the response of maize (Zea mays L. cv. Anjou 256)to a simultaneous, but separated supply of ammonium and nitrate(localized supply, LS). A split-root system was used to supplyhalf of the roots with ammonium and the other half with nitrate.A homogeneously distributed supply of both nitrogen forms (HS)was the control treatment. Seedlings were grown for 12 d fromthe two-leaf to the three-leaf stage in hydroponics at threepH levels (4, 5·5 and 7). The total N concentration was3 mol m^-3. The split-root system was established by removingthe seminal root system and using only four nodal roots perplant. Total root length and root surface area were recordedautomatically with a modified Delta- T area meter. Other morphologicalroot traits (such as main axis length and diameter, number,density, and length of laterals) were recorded manually. Uptakeof ammonium and nitrate was measured by the depletion of thenutrient solution. As compared with LS, HS was superior in shootand root DM, total root length and root surface area, ammoniumand nitrate uptake and shoot nitrogen concentration, irrespectiveof pH level. This indicates that, also under field conditions,mixed ammonium and nitrate fertilization is only beneficialto plant growth if both N forms are evenly distributed in thesoil. At both HS and LS, ascending pH increased the ammonium:nitrateuptake ratio. At LS, declining pH induced a considerable shiftin the distribution of root DM, root length, and root surfacearea the nitrate-fed compartment.Copyright 1993, 1999 AcademicPress Maize, Zea may L., ammonium, nitrate, pH, root morphology, split-root     

Fluorescent detection-polymerase chain reaction (FD-PCR) assay on microwell plates as a screening test for salmonellas in foods

This study evaluates a polymerase chain reaction assay coupled with a fluorescent detection in microwell plates for salmonellas in food samples. Chelex 100-extracted cultures and bulk and processed food samples were used as templates for a PCR assay in microwell plates, with a primer pair that amplifies a 206 bp segment of IS 200 . The PCR products were then denatured by heat and transferred to CovaLink NH plates (Nunc) to which capture oligonucleotides were covalently bound. Hybridization was performed for 1 h at 55°C, the microwells were washed and an alkaline phosphatase-labelled probe, complementary of an internal sequence of the PCR product, was added. After stringent washes, 100 μl of 1 mmol 1^-1 AttoPhos^TM (JBL Scientific) was then added to the wells and the fluorescence measurement system (Millipore). The level of detection of the assay was as low as 1–10 cfu. A total of 172 food samples were tested, both by culture and FD-PCR. Of these 53 were culture positive and 119 culture negative. The sensitivity of the FD-PCR assay was 100% and the specificity was 90.1%. Positive and negative predictive values were 82.8 and 100%, respectively. Based on the results obtained in this study it appears that the FD-PCR. assay described here can be useful to screen a large number of food samples for contamination by salmonellas.