Influence of oestradiol on protein expression and methionine utilization during morphogenesis of Paracoccidioides brasiliensis

The temporal sequence of cytosolic protein expression during phase transition of Paracoccidioides brasiliensis was examined. Electrophoretic analysis of cytosol proteins by one-dimensional SDS-PAGE revealed numerous differences between the mycelial and yeast forms as well as alterations induced by 17 beta-oestradiol. Using either protein staining or fluorography of [35S]methionine-labelled proteins 30 phase-specific bands were detected, 12 mycelial-associated bands (range 30 to 140 kDa) and 18 yeast-associated bands (range 22 to 127 kDa). In cells undergoing mycelial to yeast transition after a shift from 25 degrees C to 37 degrees C, the protein patterns showed a temporal progression toward the yeast profile with the accumulation of yeast bands prior to observable morphogenesis. Five novel protein bands (range 23 to 50 kDa) were detected by silver staining during transition. Treatment of temperature-shifted mycelial cultures with 2.6 x 10(-7) M-oestradiol altered observed profiles; 4 of 12 mycelial-associated bands were maintained whereas the appearance of the 5 novel transition bands and 9 of 18 yeast-associated bands was blocked or delayed. Analysis of [35S]methionine-labelled proteins revealed that oestradiol induced label uptake by mycelial cells, blocked the synthesis of a 92 kDa yeast-specific band 72 h into transition, and diminished label incorporation 120 h into transition. In conjunction with these steroid-induced alterations of protein expression, little or no morphological transformation occurred. These results support our hypothesis that, analogous to mammalian steroid receptor action, the functional responses of P. brasiliensis to oestradiol are related to regulation of protein expression, presumably mediated via a specific binding protein-ligand complex.     

Contraception in mares heteroimmunized with pig zonae pellucidae

Ten fertile feral mares and 6 domestic horses (4 fertile mares, 1 infertile mare, 1 gelding) were immunized with heat-solubilized pig zonae pellucidae by 4 injections equivalent to 2000 or 5000 zonae each at 2-4-week intervals and a booster injection of 20,000 zonae 6-10 months after the last of the initial inoculations. The immune response was reflected by high antibody levels as measured by an enzyme-linked immunosorbent assay (ELISA) using immobilized pig zona antigen. In-vivo inhibition of fertility occurred in 12 (86%) of the 14 fertile mares studied and persisted for a minimum of 7 months. Repeated mating of the fertile domestic mares resulted in conception when anti-pig zona antibody concentrations had decreased from initial peak absorbance ratios (greater than 1.0) to relatively lower levels (0.64 or less with one exception). An indirect immunofluorescence assay, revealed a considerably lower cross-reactive antibody titre with horse oocytes as compared to pig oocytes. Clinical, endocrinological and histological analyses of the ovaries and their function following regained fertility after immunization revealed no abnormalities. One mare remained infertile.     

The precise radioimmunoassay of adenosine: minimization of sample collection artifacts and immunocrossreactivity.

Anti-adenosine antibodies were produced in rabbits immunized with N6-carboxymethyladenosine conjugated to methyl albumin. 125I-N6-Aminobenzyladenosine was synthesized and used as a high-specific-activity, high-affinity ligand. A radioimmunoassay (RIA) was developed that can detect 6.25 nM (312.5 fmol) of underivatized adenosine and cross-reacts less than 0.02% with adenine nucleotides and guanosine and not at all with 1 mM inosine. The sensitivity of the RIA can be increased to a detection limit of 0.125 nM (6.25 fmol) by derivitizing samples with benzyl bromide to form N6-benzyladenosine. The assay was adapted to an automated RIA procedure. Assay precision was increased by: (i) inhibiting slight adenosine deaminase activity present in anti-sera; (ii) treating buffers and albumin used in the RIA with charcoal to remove contaminating adenosine; and (iii) correcting for a small but variable component of immunoreactivity not attributable to adenosine. A second antibody prepared with a 2',3'-disuccinyladenosine-albumin conjugate was also found to detect some non-adenosine-mediated immunoreactivity in plasma samples. Immunointerference in human plasma was eliminated in samples treated with ZnSO4/Ba(OH)2 or partially purified over C18 Sep Paks to remove nucleotides and assayed after sample benzylation or succinylation. Human blood was mixed with a novel "stop" solution that was optimized to inhibit adenosine formation from AMP by greater than 99% and to inhibit adenosine uptake into red cells and degradation by greater than 94%. Human plasma/stop solution was assayed by RIA and HPLC with equivalent results.     

Gene-culture coevolutionary theory

Gene-culture coevolutionary theory is a branch of theoretical population genetics that models the transmission of genes and cultural traits from one generation to the next, exploring how they interact. These models have been employed to examine the adaptive advantages of learning and culture, to investigate the forces of cultural change, to partition the variance in complex human behavioral and personality traits, and to address specific cases in human evolution in which there is an interaction between genes and culture.     

Characterization of a calcium/calmodulin-dependent protein kinase homolog from maize roots showing light-regulated gravitropism

Roots of many species respond to gravity (gravitropism) and grow downward only if illuminated. This light-regulated root gravitropism is phytochrome-dependent, mediated by calcium, and inhibited by KN-93, a specific inhibitor of calcium/calmodulin-dependent protein kinase II (CaMK II). A cDNA encoding MCK1, a maize homolog of mammalian CaMK, has been isolated from roots of maize (Zea mays L.). The MCK1 gene is expressed in root tips, the site of perception for both light and gravity. Using the [³⁵S]CaM gel-overlay assay we showed that calmodulin-binding activity of the MCK1 is abolished by 50 M KN-93, but binding is not affected by 5 M KN-93, paralleling physiological findings that light-regulated root gravitropism is inhibited by 50 M KN-93, but not by 5 M KN-93. KN-93 inhibits light-regulated gravitropism by interrupting transduction of the light signal, not light perception, suggesting that MCK1 may play a role in transducing light. This is the first report suggesting a physiological function for a CaMK homolog in light signal transduction.Abbreviations CaM calmodulin - CaMK (II) Ca²⁺/calmodulin-dependent protein kinase (II) - CBP CaM-binding protein - CDPK Ca²⁺-dependent protein kinase - MCK1 maize homolog of mamalian CaMK This work is supported by the National Aeronautics and Space Administration grant No: NAGW 238.     

High-performance liquid chromatographic determination of iobitridol in plasma, urine and bile

Iobitridol is a new non-ionic, low-osmolality contrast medium for urography and angiography. We have developed a method for determining iobitridol in body fluids using high-performance liquid chromatography with ultraviolet detection. The method, which is specific and reproducible, does not require an internal standard. Determinations can be carried out in body fluids against a set of standards in ethanol. The method was validated for the quantification of iobitridol in biological samples obtained during pharmacokinetic studies.     

Bi-directional inflorescence development inArabidopsis thaliana: Acropetal initiation of flowers and basipetal initiation of paraclades

In this study we investigated Arabidopsis thaliana (L.) Heynh. inflorescence development by characterizing morphological changes at the shoot apex during the transition to flowering. Sixteen-hour photoperiods were used to synchronously induce flowering in vegetative plants grown for 30 d in non-inductive 8-h photoperiods. During the first inductive cycle, the shoot apical meristem ceased producing leaf primordia and began to produce flower primordia. The differentiation of paraclades (axillary flowering shoots), however, did not occur until after the initiation of multiple flower primordia from the shoot apical meristem. Paraclades were produced by the basipetal activation of buds from the axils of leaf primordia which had been initiated prior to photoperiodic induction. Concurrent with the activation of paraclades was the partial suppression of paraclade-associated leaf primordia, which became bract leaves. The suppression of bract-leaf primordia and the abrupt initiation of flower primordia during the first inductive photoperiod is indicative of a single phase change during the transition to flowering in photoperiodically induced Arabidopsis. Morphogenetic changes characteristic of the transition to flowering in plants grown continuously in 16-h photoperiods were qualitatively equivalent to the changes observed in plants which were photoperiodically induced after 30 d. These results suggest that Arabidopsis has only two phases of development, a vegetative phase and a reproductive phase; and that the production of flower primordia, the differentiation of paraclades from the axils of pre-existing leaf primordia and the elongation of internodes all occur during the reproductive phase.     

Helicobacter pylori and peptic ulcer disease.

Medical therapy for duodenal or gastric ulcer disease has traditionally involved gastric acid antisecretory therapy for 4 to 8 weeks to promote initial healing and indefinitely to prevent recurrences of ulcer. The discovery of Helicobacter pylori in most patients with peptic ulcer disease has led to a change in this approach. Therapy designed to eradicate H pylori may facilitate ulcer healing with acid antisecretory agents and, more important, may greatly reduce the incidence of ulcer recurrence, obviating the need for maintenance antisecretory therapy. Regimens designed to eradicate H pylori are difficult to comply with, however, and are associated with adverse effects in some patients. In this article we review the diagnosis and treatment of H pylori infection in patients with peptic ulcer disease and make recommendations regarding the use of conventional ulcer therapies and therapies designed to eradicate H pylori.     

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