Regulation of tyrosinase during the vegetative and sexual life cycles of Neurospora crassa

Tyrosinase derepression in Neurospora mycelia grown in Vogel medium, submitted to starvation in phosphate buffer 0.1 M, pH 6.0, was abolished by exogenous magnesium sulfate. This effect seemed to be caused by the sulfate ion itself and not by a sulfate-derivative. Sulfate repression required protein synthesis, thus suggesting the involvement of a specific gene product mediating sulfate repression. Cultures made in Westergaard and Mitchell crossing medium became competent for sexual development and could be stimulated to form tyrosinase either by mating or starvation. In that case the enzyme derepression was insensitive to the sulfate effect. The possible existence of a positive mechanism for the control of tyrosinase activity during sexual development is suggested.This work is a part of two theses, by Rolf Alexander Prade and Angela Kaysel Cruz submitted to the Departments of Biochemistry and Physiology, respectively, of the School of Medicine of Ribeirão Preto in partial fulfillments of the requirements for the Master Degree.     

Persistent infection of cells in culture by measles virus. I. Development and characteristics of HeLa sublines persistently infected with complete virus

Rustigian, Robert (Tufts University School of Medicine, Boston, Mass.). Persistent infection of cells in culture by measles virus. I. Development and characteristics of HeLa sublines persistently infected with complete virus. J. Bacteriol. 92:1792-1804. 1966.-After the development of marked cytopathic effects in HeLa cultures infected with the Edmonston strain of measles virus, renewed cell growth occurred, and HeLa sublines persistently infected with measles virus were obtained. Persistent infection has occurred in a large fraction of the cells of infected clonal lines for more than 300 to 500 cell generations during a period of 6 years. One mechanism by means of which infection was maintained in the clonal lines is transmission of virus or viral subunits from cell to cell at division. Continued subculture of the persistently infected populations resulted in the virtual disappearance of cytopathic effects, a marked decrease in the amount of extracellular virus, and alterations in the cytopathogenicity of virus recovered from persistently infected populations. The intracellular virus-host cell events in late passages of the infected clonal lines appeared to be similar to those in cells of primary infected cultures at early stages of infection, as judged by the pattern of viral immunofluorescence and the very low incidence of cells with intranuclear inclusion bodies. Cultures of the persistently infected clonal lines were highly resistant to super infection by parent Edmonston virus. Cultures of one of these clonal lines were just as susceptible as normal HeLa cultures to vaccinia, herpes simplex, and polio type 2 viruses, and a simian agent, with a possible low degree of resistance to the simian agent.     

School-based health promotion: the physician as advocate

At the August 1995 meeting of the General Council of the CMA, a resolution supporting school-based health promotion (comprehensive School Health) was adopted. This article briefly reviews the research supporting this integrated approach to school and community programs, applies the recommended approach to reducing tobacco use and outlines a role for physicians in promoting Comprehensive School Health in their communities.     

Gene regulatory network inference and validation using relative change ratio analysis and time-delayed dynamic Bayesian network

The Dialogue for Reverse Engineering Assessments and Methods (DREAM) project was initiated in 2006 as a community-wide effort for the development of network inference challenges for rigorous assessment of reverse engineering methods for biological networks. We participated in the in silico network inference challenge of DREAM3 in 2008. Here we report the details of our approach and its performance on the synthetic challenge datasets. In our methodology, we first developed a model called relative change ratio (RCR), which took advantage of the heterozygous knockdown data and null-mutant knockout data provided by the challenge, in order to identify the potential regulators for the genes. With this information, a time-delayed dynamic Bayesian network (TDBN) approach was then used to infer gene regulatory networks from time series trajectory datasets. Our approach considerably reduced the searching space of TDBN; hence, it gained a much higher efficiency and accuracy. The networks predicted using our approach were evaluated comparatively along with 29 other submissions by two metrics (area under the ROC curve and area under the precision-recall curve). The overall performance of our approach ranked the second among all participating teams.     

Bilingual Education: A Dialogue with the Bakhtin Circle.

Bilingual Education: A Dialogue with the Bakhtin Circle. Marcia Moraes. Albany: State University of New York Press, 1996. 159 pp.     

Dialogue and the Interpretation of Illness. Conversations in a Cameroon Village

Dialogue and the Interpretation of Illness. Conversations in a Cameroon Village. Robert Pool. Oxford, UK & Providence, RI: Berg Publishers, 1994 (cloth and paper). xii + 286 pp.     

Decolonizing Knowledge: From Development to Dialogue

Decolonizing Knowledge: From Development to Dialogue. Frederique Apffel-Marglin and Stephen A. Marglin. eds. Oxford, England: Clarendon Press, 1996. 398 pp.     

L��h��ritage de l����cole de Boston

The Boston School was a school of thought that gave birth, in the 1890s in the USA, to psychology as an academic discipline and as a practising profession. Some of the ideas and therapeutic attitudes of the school??s members will be examined from a historical perspective. The approach will be compared briefly with that of current psychological practice in France. The aim of this comparison is to get an overview of the development of psychology from its initial progress in the USA to current practice in France and from earlier European influences to the present American contribution. Thus, we can explore the legacy of the Boston School and some more recent potential influences. In comparison with modern psychologists how did the members of the Boston School regard psychology, mental life and psychotherapies?     

Effects of Fatty Acid Substitution on the Release of Enzymes by Osmotic Shock

The release of enzymes by osmotic shock from Escherichia coli strain 30E, an unsaturated fatty acid auxotroph, was examined in culture supplemented with either cis- or trans-unsaturated fatty acids. Cultures grown in oleate-supplemented medium release a large fraction of the total cyclic phosphodiesterase, acid hexose phosphatase, and 5'-nucleotidase following osmotic shock. Cultures grown in elaidate-supplemented medium release much less of these same enzymes after shock treatment. Cultures grown with either supplementation show total release of these enzymes upon conversion to spheroplasts, demonstrating that the enzymes are in the periplasmic space in both cases. Cultures grown with either oleate or elaidate as fatty acid source were washed and suspended in medium containing the other isomer. The change from oleate to elaidate resulted in a rapid decrease in ability of the cells to release the three enzymes after osmotic shock so that within a 25% increase in cell mass the culture responded to osmotic shock as would a culture grown overnight in elaidate-supplemented medium. The reverse experiment resulted in a gradual increase in the ability of the cells to respond to osmotic shock. The outer membrane of E. coli is altered by the incorporation of elaidate, as indicated by electron microscopic data.     

Teamwork: Improved eQTL Mapping Using Combinations of Machine Learning Methods

Expression quantitative trait loci (eQTL) mapping is a widely used technique to uncover regulatory relationships between genes. A range of methodologies have been developed to map links between expression traits and genotypes. The DREAM (Dialogue on Reverse Engineering Assessments and Methods) initiative is a community project to objectively assess the relative performance of different computational approaches for solving specific systems biology problems. The goal of one of the DREAM5 challenges was to reverse-engineer genetic interaction networks from synthetic genetic variation and gene expression data, which simulates the problem of eQTL mapping. In this framework, we proposed an approach whose originality resides in the use of a combination of existing machine learning algorithms (committee). Although it was not the best performer, this method was by far the most precise on average. After the competition, we continued in this direction by evaluating other committees using the DREAM5 data and developed a method that relies on Random Forests and LASSO. It achieved a much higher average precision than the DREAM best performer at the cost of slightly lower average sensitivity.     

Production of extracellular phytase fromAspergillus ficuum on starch media

Summary Cultures grown on Hylon corn starch media produced the highest levels of phytase. Phospholipid extraction of Hylon starch did not change its effectiveness as a substrate. Cultures grown on phosphorylated dextrins from Hylon corn starch produced equivalent amounts of phytase.     

Leukocytes and interferon in the host response to viral infections. II. Enhanced interferon response of leukocytes from immune animals

Glasgow, Lowell A. (University of Rochester School of Medicine and Dentistry, Rochester, N.Y.). Leukocytes and interferon in the host response to viral infections. II. Enhanced interferon response of leukocytes from immune animals. J. Bacteriol. 91:2185-2191. 1966.-The production of interferon was studied under in vitro conditions in peritoneal leukocytes or macrophages from mice immunized with Chikungunya virus (CV). Cultures of leukocytes obtained from animals immune to CV produced 2- to 10-fold greater amounts of interferon when exposed to an inoculum of CV than similar cell preparations from nonimmune, control animals. The viral inhibitor produced in increased quantity by CV-immune leukocytes had the biological and biochemical properties of interferon. The enhanced interferon production was inhibited by actinomycin D. This response of immune leukocytes was specific, and was initiated only by CV; it was not observed in leukocytes from animals immunized against other viruses which were challenged with CV. The presence of neutralizing antibody could not be related to this response. The observed increase in interferon production was not dependent upon an enhanced virus uptake. The data are presented as a possible new dimension of the "immune response" and may suggest a mechanism for the phenomenon of "tissue immunity."     

Maintenance of in vitro cultures of Babesia divergens and Babesia major at low temperatures

Cultures of erythrocytes parasitized by Babesia divergens and Babesia major were stored in medium cooled to 4 degrees C for up to 8 weeks. There was a marked decrease in parasitaemia and an increase in the number of free extra-erythrocytic, unagglutinated merozoites, during the cooled period. Cultures stored in this way and returned to 38 degrees C resumed growth, with or without sub-culture. At the low temperature, only one sub-culture is required per week.     

Long-term maintenance of primary myogenic cultures on a reconstituted basement membrane

Summary We describe a simple technique for maintaining highly contractile long-term chicken myogenic cultures on Matrigel, a gel composed of basement membrane components extracted from the Engelbreth-Holm-Swarm mouse tumor. Cultures grown on Matrigel consist of three-dimensional multilayers of cylindrical, contracting myotubes which endure for at least 60 d without myotube detachment. A Matrigel substrate increases the initial plating efficiency but does not effect cell proliferation. Large-scale differentiation in cultures maintained on Matrigel is delayed by 1 to 2 d, compared to cultures grown on gelatin-coated dishes. Long-term maintenance on Matrigel also results in increased expression of the neonatal and adult fast myosin heavy chain isoforms. Culturing of cells on a Matrigel substrate could thus facilitate the study of later events of in vitro myogenesis. This work was supported by grants to Z. Y.-R. from the American Heart Association Washington Affiliate, the University of Washington Graduate School Research Fund, and the National Institutes of Health, Bethesda, MD (AR39677). R. S. H. was supported by a Predoctoral Developmental Biology Training Grant from the National Institutes of Health (HD07183-10). Note Added in Proof Strohman et al. (31) have recently reported on the expression of neonatal and adult isoforms of fast myosin heavy chain in chicken myogenic cultures maintained on flexible membranes.     

Concomitant quantification of targeted drug delivery and biological response in individual cells

Targeted therapies result in heterogeneous drug delivery, often with highly variable drug uptake in the targeted cells and significant numbers of cells that are essentially untargeted. However both the variably targeted cells and neighboring bystander cells may respond to the treatment. Using ionizing radiation as an example of a targeted therapeutic agent, we describe a quantitative immunofluorescence-based approach for concomitant quantification of exposure and measurement of biological responses in both targeted and bystander cells. Cultures of human skin fibroblasts are co-pulse-labeled with 3H-deoxycytidine (3H-dC) and bromodeoxyuridine (BrdU). The labeled cells, identified by BrdU immunofluorescence, are internally irradiated by low-energy beta-particles emitted by incorporated 3H-dC. BrdU immunofluorescence intensity is proportional to radioactivity incorporated and, therefore, to radiation dose rate. Cell-cycle arrest in G2 is measured in labeled cells as function of dose rate. Stress responses in bystander cells, indicated by a G1 checkpoint, are concomitantly measured with a flow cytometric-cumulative labeling index (FCM-CLI) assay. The overall approach presented herein may be useful in the context of evaluating responses to targeted drug delivery.     

Paramoeba sp. (Amoebida, Paramoebidae) as the possible causative agent of sea urchin mass mortality in Nova Scotia

An amoeba resembling Paramoeba has been cultured from tissues of diseased sea urchins. Cultures containing the amoeba produced signs of the disease when injected into the coelom of healthy urchins. Control treatments lacking the amoeba did not cause the disease. The amoeba was cultured from radial nerve fragments and seen in tissue sections from experimentally infected urchins. Cultures of the amoeba from these experimentally infected urchins reproduced the disease in healthy urchins by both injection and water-borne routes. These observations suggest involvement of Paramoeba in recent mass mortalities of sea urchins in Nova Scotia.     

Characterization of primary and secondary forms of Xenorhabdus luminescens strain Hm

Abstract Cultures of the primary form of Xenorhabdus luminescens strain Hm gave rise to secondary forms after prolonged static incubation in two broth media. The secondary forms were deficient in pigmentation and extracellular antibiotic, protease and lipase activities, and were about 100-fold less luminous than the primary form in vivo. Secondary forms isolated on two separate occasions from two different media were identical in their deficiencies. Cultures of the secondary form in defined broth media produced no detectable secondary metabolites, unlike the primary form, and grew more rapidly than the primary form. A protocol for screening primary cultures of X. luminescens for secondary forms is presented.     

Detection of mutation in transgenic CHO cells using green fluorescent protein as a reporter

A novel approach was developed for rapidly estimating the frequency of specific mutations in genetically engineered Chinese hamster ovary (CHO) cells. We designed double-transgenic CHO cell lines that contain a transgene consisting of the sequence coding for green fluorescent protein under the control of a tetracycline (Tet) responsive promoter and a second transgene coding for the constitutively expressed Tet repressor. Cultures of these CHO cells were treated with gamma-radiation, N-methyl-N-nitrosourea or methyl methanesulfonate, and the fluorescence of individual cells from both control and treated cultures was measured by flow cytometry. The treatments increased the number of highly fluorescent cells, those with presumed mutations in the Tet-repressor gene. Mutant cells from gamma-radiation-exposed cultures were isolated by fluorescence-activated cell sorting, cultured, and individual clones expanded. A PCR-based analysis indicated that the highly fluorescent expanded cells had lost the transgene coding for the Tet repressor, suggesting that the system mainly detects large genetic alterations. A similar approach may be useful for making high-throughput in vivo models for mutation detection.     

Cultivation at 37°C of a Trypanosome (Trypanosoma theileri) from Cows with Depressed Milk Production.

SYNOPSIS. Cultures of Trypanosoma theileri were obtained at 36° and at 37.5°C. in a blood-lysate medium inoculated with blood from three dairy cows showing subnormal milk production. The organisms were first seen after 4 days in the first subculture, reached a maximum number of about 500,000 per ml. on the 4th day of the second subculture, and attained about this same number on the 4th day of subsequent transfers. Crithidial forms predominated but trypanosomes of the blood-stream type were also numerous. Cultures were not obtained from cows with normal milk production. The infected cows, although free from helminth parasites, showed a marked eosinophilia.