Stimulation of bacterial cytokinesis by bacteriophage predation

Bacteriophage (phage) are ubiquitous in the water column and in the sediments of most natural water. Because of their colloidal nature, they can either aggregate into clumps large enough to settle into the sediment or departing upon the physiochemical conditions or disassociate and reenter the water column. About 80% of the bacterial strains isolated from New River sediment have a virulent phage that can be isolated with them.Liquid cultures of a strain of Pseudomonas aeruginosa isolated from the New River along with its phage were set up. One was infected with the virulent phage and another kept as a control. Daily counts were made of bacterial numbers. After 10 days the control culture was infected and counted for 3 more days.Both cultures divided exponentially at first. The infected culture continued to divide at about half the initial rate. The uninfected culture nearly ceased division, but when phage were added it quickly began to divide.The virulent phage infection clearly stimulated host division. The effect was to establish itself as an endemic infection which did not outpace its host's division rate. Further, the enhanced division rate may act to increase the host's share of available nutrients and benefit its competitive position in the system.     

Crystallization of the A-domain of the mannitol transport protein enzyme IImtl.

The A-domain of the mannitol transport protein enzyme IImtl from Escherichia coli (relative molecular mass 16,300) was crystallized, both at room temperature and 4 degrees C, from 40% polyethylene glycol 6000 (pH 8.5 to 9.0) using the hanging-drop method of vapour diffusion. The crystals have the monoclinic space group P2(1), with unit cell dimensions a = 54.0 A, b = 67.0 A, c = 80.9 A and beta = 100.8 degrees. They diffract to 2.6 A resolution. A self-rotation function and self-Patterson suggest that there are four molecules in the asymmetric unit showing mmm symmetry.     

Interaction of phosphatidylinositol 3-kinase-associated p85 with epidermal growth factor and platelet-derived growth factor receptors.

One of the immediate cellular responses to stimulation by various growth factors is the activation of a phosphatidylinositol (PI) 3-kinase. We recently cloned the 85-kDa subunit of PI 3-kinase (p85) from a lambda gt11 expression library, using the tyrosine-phosphorylated carboxy terminus of the epidermal growth factor (EGF) receptor as a probe (E. Y. Skolnik, B. Margolis, M. Mohammadi, E. Lowenstein, R. Fischer, A. Drepps, A. Ullrich, and J. Schlessinger, Cell 65:83-90, 1991). In this study, we have examined the association of p85 with EGF and platelet-derived growth factor (PDGF) receptors and the tyrosine phosphorylation of p85 in 3T3 (HER14) cells in response to EGF and PDGF treatment. Treatment of cells with EGF or PDGF markedly increased the amount of p85 associated with EGF and PDGF receptors. Binding assays with glutathione S-transferase (GST) fusion proteins demonstrated that either Src homology region 2 (SH2) domain of p85 is sufficient for binding to EGF and PDGF receptors and that receptor tyrosine autophosphorylation is required for binding. Binding of a GST fusion protein expressing the N-terminal SH2 domain of p85 (GST-N-SH2) to EGF and PDGF receptors was half-maximally inhibited by 2 and 24 mM phosphotyrosine (P-Tyr), respectively, suggesting that the N-SH2 domain interacts more stably with PDGF receptors than with EGF receptors. The amount of receptor-p85 complex detected in HER14 cells treated with EGF or PDGF. Growth factor treatment also increased the amount of p85 found in anti-PDGF-treated HER14 cells, suggesting that the vast majority of p85 in the anti-P-Tyr fraction is receptor associated but not phosphorylated on tyrosine residues. Only upon transient overexpression of p85 and PDGF receptor did p85 become tyrosine phosphorylated. These are consistent with the hypothesis that p85 functions as an adaptor molecule that targets PI 3-kinase to activated growth factor receptors.     

Comparative effects of trichothecene mycotoxins on bovine platelet function: Acetyl T-2 toxin,a more potent inhibitor than T-2 toxin

The effects of the trichothecene mycotoxins (acetyl T-2 toxin, T-2 toxin, HT-2 toxin, palmityl T-2 toxin, diacetoxyscirpenol (DAS), deoxynivalenol (DON), and T-2 tetraol) on bovine platelet function were examined in homologous plasma stimulated with platelet activating factor (PAF). The mycotoxins inhibited platelet function with the following order of potency: acetyl T-2 toxin > palmityl T-2 toxin = DAS > HT-2 toxin = T-2 toxin. While T-2 tetraol was completely ineffective as an inhibitor, DON exhibited minimal inhibitory activity at concentrations above 10×10^?4M. The stability of the platelet aggregates formed was significantly reduced in all mycotoxin treated platelets compared to that of the untreated PAF controls. It is suggested that the increased sensitivity of PAF stimulated bovine platelets to the more lipophilic mycotoxins may be related to their more efficient partitioning into the platelet membrane compared to the more hydrophilic compounds.     

Deoxyribonucleotide biosynthesis in yeast: assay and properties of ribonucleotide reductase in permeabilized Saccharomyces cerevisiae cells

Yeast cells permeabilized by freeze-thaw cycles in a sorbitol-containing medium provide an experimentally favorable system for the study of ribonucleotide reduction in a small number of cells or in mutant strains. Ribonucleotide reductase activities determined in such cells are about twice those found in cell extracts but properties of the enzyme, except pH optimum, are closely comparable in both assay procedures. In contrast with other organisms, the activities measured in permeabilized cells from both diploid or haploid strains exceed the demand for deoxyribonucleotide formation during replication of the yeast genome. The method has been applied to yeast cultures growing in the presence of the ribonucleotide reductase inhibitor hydroxyurea and a twofold increase of enzyme activity has been established in such cells. On the other hand, analysis of a series of hus mutants, selected for hydroxyurea sensitivity in the laboratory of Singer and Johnston did not reveal obvious alterations of the enzyme vs the parental strains, suggesting that the hus phenotype may be due to lesions other than in ribonucleotide reductase.     

Reproduction and social rank in female stumptail Macaques (Macaca arctoides)

Reproductive physiology was studied in female stumptail macaques. Initially the monkeys were housed indoors (individually and in small groups) and later as one large (92 individuals) social group in an outdoor cage. Most data were collected during the 4-year outdoor period. Plasma progesterone determination in blood samples taken at weekly intervals allowed estimation of ovulation and conception dates. The age at first ovulation (X =3.73 years) was positively correlated with body weight at 3 years of age. The average age at first birth was 4.90 years. Gestation lengths averaged 176.6 days. Following a live birth ovulations returned after a mean interval of 11 months but following an abortion or still birth this interval was 1 month. Usually a number of ovulatory cycles (X =2.37) preceded a conception. Interbirth intervals (IBIs) in the outdoor cage (X =619.4 days) were significantly longer than IBIs during the indoor period (X =523.1), because indoors the infants were weaned at the age of 7 months, while outdoors weaning occurred more naturally. IBIs following abortions or still births (X =291.9 days) were significantly shorter than IBIs following live births. Age at first ovulation, age at first birth, IBIs, and infant production rates were not correlated with dominance rank. Ovarian cycle lengths (X =30.2 days, mode = 28 days) were comparable to previously reported data from laboratory-housed stumptails. No systematic seasonal fluctuations were found in the onset of sexual maturity, in ovarian cycle lengths, in copulation frequencies, and in distribution of births.     

Binding of chondroitin sulfate, dermatan sulfate and fat-storing cell-derived proteoglycans to rat hepatocytes

1. The interaction of isolated rat hepatocytes with exogenous 3H-labeled chondroitin-4-sulfate and dermatan sulfate and with biosynthetically 35S-labeled proteoglycans secreted by cultured rat liver fat-storing cells has been studied. 2. All ligands are bound by hepatocytes in a concentration-dependent manner. Scatchard-plot analysis of the data revealed the existence of high- and low-affinity binding modes. 3. The cell-bound exogenous [3H]glycosaminoglycans could be displaced by each unlabeled ligand and by heparin, whereas displacement of the endogenous material was less effective. 4. Binding of all ligands to hepatocytes increased with time. For the exogenous glycosaminoglycans the two- to threefold amount was retained at 37 degrees C as compared to 4 degrees C; it was markedly reduced by pretreatment of the cells with trypsin. 5. Degradation of the exogenous ligands could be detected neither for the cell-bound fraction nor for the free glycosaminoglycans in the culture medium. 6. The binding of the ligands to hepatocytes is viewed as a cell-matrix interaction. Its possible pathobiochemical relevance in liver fibrosis or neoplasia is discussed.     

Enhancement of translational efficiency by the Escherichia coli atpE translational initiation region: its fusion with two human genes

The cDNA sequences encoding mature human interleukin 2 (IL2) and beta-interferon (INF beta), respectively, were fused with various translational initiation regions and inserted into two different types of expression vector. The relative levels of expression of the two genes and the functional stability of their respective mRNAs were examined in vivo in Escherichia coli hosts. The addition of the 30-bp sequence, found immediately upstream of the E. coli atpE gene Shine-Dalgarno (SD) sequence, to the translational initiation regions of IL2 and INF beta increased the expression of both these genes by a factor of 6-10. Thus this sequence, which naturally acts within the E. coli atp operon to enhance the translational initiation frequency of the atpE gene, can increase the expression of other genes in E. coli. It may exemplify a specific type of recognition signal for the E. coli translational apparatus.     

β-adrenoceptors in human tracheal smooth muscle: characteristics of binding and relaxation

Specific binding of [¹²⁵I]-(−)-cyanopindolol to human tracheal smooth muscle membranes was saturable, stereo-selective and of high affinity (K_d=5.3±0.9 pmol/l and R_T=78±7fmol/g tissue). The β₁-selective antagonists atenolol and LK 203-030 inhibited specific [¹²⁵I]-(−)-cyanopindolol binding according to a one binding site model with low affinity in nearly all subjects, pointing to a homogeneous β₂-adrenoceptor population. In one subject using LK 203-030 a small β-adrenoceptor subpopulation could be demonstrated. The beta-mimetics isoprenaline, fenoterol, salbutamol and terbutaline recognized high and low affinity agonist binding sites. Isoprenaline's pK_H- and pK_L- values for the high and low affinity sites were 8.0±0.2 and 5.9±0.3 respectively. In functional experiments isoprenaline relaxed tracheal smooth muscle strips having intrinsic tone with a pD₂-value of 6.63±0.19.     

A mutation in a case of early onset narcolepsy and a generalized absence of hypocretin peptides in human narcoleptic brains

We explored the role of hypocretins in human narcolepsy through histopathology of six narcolepsy brains and mutation screening of Hcrt, Hcrtr1 and Hcrtr2 in 74 patients of various human leukocyte antigen and family history status. One Hcrt mutation, impairing peptide trafficking and processing, was found in a single case with early onset narcolepsy. In situ hybridization of the perifornical area and peptide radioimmunoassays indicated global loss of hypocretins, without gliosis or signs of inflammation in all human cases examined. Although hypocretin loci do not contribute significantly to genetic predisposition, most cases of human narcolepsy are associated with a deficient hypocretin system.