Effects of angiogenin on granulosa and theca cell function in cattle

Angiogenin is a member of the ribonuclease A superfamily of proteins that has been implicated in stimulating angiogenesis but whether angiogenin can directly affect ovarian granulosa or theca cell function is unknown. Therefore, the objective of these studies was to determine the effect of angiogenin on proliferation and steroidogenesis of bovine granulosa and theca cells. In experiments 1 and 2, granulosa cells from small (1 to 5 mm diameter) follicles and theca cells from large (8 to 22 mm diameter) follicles were cultured to evaluate the dose-response effect of recombinant human angiogenin on steroidogenesis. At 30 and 100 ng/ml, angiogenin inhibited (P<0.05) granulosa cell progesterone production and theca cell androstenedione production but did not affect (P>0.10) granulosa cell estradiol production or theca cell progesterone production, and did not affect numbers of granulosa or theca cells. In experiments 3 and 4, granulosa and theca cells from both small and large follicles were cultured with 300 ng/ml of angiogenin to determine if size of follicle influenced responses to angiogenin. At 300 ng/ml, angiogenin increased large follicle granulosa cell proliferation but decreased small follicle granulosa cell progesterone and estradiol production and large follicle theca cell progesterone production. In experiments 5 and 6, angiogenin stimulated (P<0.05) proliferation and DNA synthesis in large follicle granulosa cells. In experiment 7, 300 ng/ml of angiogenin increased (P<0.05) CYP19A1 messenger RNA (mRNA) abundance in granulosa cells but did not affect CYP11A1 mRNA abundance in granulosa or theca cells and did not affect CYP17A1 mRNA abundance in theca cells. We conclude that angiogenin appears to target both granulosa and theca cells in cattle, but additional research is needed to further understand the mechanism of action of angiogenin in granulosa and theca cells, as well as its precise role in folliculogenesis.     

Effect of cocaine and morphine on neutral endopeptidase activity of human peripheral blood mononuclear cells cultured with lectins

We have tested the effect of alkaloids (cocaine, morphine) and enkephalins on neutral endopeptidase of peripheral blood mononuclear cells activated by lectins. When treated with concanavalin A and cocaine, peripheral blood mononuclear cells showed an enhanced activity (+110 per cent) of the membrane neutral endopeptidase, which was not related to the expression of the common acute lymphoblastic leukemia antigen at the cell surface, although both molecules have the identical amino acid sequence. Phytohemagglutinin-P, morphine and synthetic enkephalins did not induce the activity of neutral endopeptidase nor the expression of common acute lymphoblastic leukemia antigen. Our findings suggested that the drugs of abuse, cocaine and morphine, affected specific membrane constituents without altering proliferation, subcellular localization of membrane enzymes or the surface immune phenotype of peripheral blood mononuclear cells.     

Actin pegs and ultrastructure of presumed sensory receptors of Beroë (Ctenophora)

Summary We have investigated the actin content and ultrastructure of two kinds of presumed sensory projections on the lip epidermis of beroid ctenophores. Transmission electron microscopy showed that conical pegs contain a large bundle of densely packed, parallel microfilaments. Rhodamine-phalloidin brightly stained the pegs, confirming that they contain filamentous actin. Epidermal cells with actin pegs also bear a single long cilium with an onion-root structure, previously described as arising from a different type of cell. The actin peg and onion-root cilium project side-by-side, defining a polarized axis of the cell which is shared by neighboring cells. The onion-root body is surrounded by a flattened membranes sac which lies immediately below the plasma membrane. The perimeter of the membrane sac is encircled by aggregates of dense material. An extra layer of dense material is found along the side of the membrane sac facing the peg; this material often makes direct contact with the adjacent actin filament bundle. Cells with actin pegs and onion-root cilia synapse onto adjacent neurites and secretory gland cells, indicating that one or both types of projections are sensory elements. Since the feeding responses of beroids are reported to depend on chemical and tactile stimuli to the lips, the cells bearing pegs and cilia may function as both mechanoreceptors and chemoreceptors, that is, as double sensory receptors.     

The dynamics of experimental arable weed communities under different management practices

Abstract. Weed communities, comprising 12 introduced species at constant starting densities and three species already present in the seed bank, were followed through three years of continuous winter wheat. The wheat and weeds were subjected to two treatments in a split-plot factorial design, organic contrasting with conventional fertilizer, and ploughing plus hand-roguing contrasting with minimum tillage plus herbicide. The minimum tilled plots developed in a uniform manner, and became dominated by very high densities of Anisantha sterilis. Agrostemma githago and Galium aparine also persisted in these plots at lower densities. The ploughed plots had a lower total density but a greater range of species. Stellaria media, Veronica persica and Avena fatua were the most common; other species occurred at lower densities. The major effect of fertilizer treatment was a greater initial increase by G. aparine on the organic, minimum tilled plots compared with the conventionally fertilized, minimum tilled plots. Species associated with minimum tillage were annuals with either no or a short term seed bank and autumn germination and rather predictable dynamics, whereas species that did well under ploughing were either spring germinating or had a persistent seed bank, implying greater annual variation in population size associated with weather conditions. There seemed no clear way to distinguish between those species which were abundant on the ploughed plots and those which were scarce under all conditions using readily accessible data.     

A recombinant foot-and-mouth disease virus antigen inhibits DNA replication and triggers the SOS response in Escherichia coli

Abstract The 3D gene of foot-and-mouth disease virus encodes the viral RNA dependent RNA polymerase, also called virus infection associated (VIA) antigen, which is the most important serological marker of virus infection. This 3D gene from a serotype Cl virus has been cloned and overexpressed in Escherichia coli under the control of the strong lambda lytic promoters. The resulting 51 kDa recombinant protein has been shown to be immunoreactive with sera from infected animals. After induction of gene expression, an immediate and dramatic arrest of cell DNA synthesis occurs, similar to that produced by genotoxic doses of the drug mitomycin C. This effect does not occur during the production of either a truncated VIA antigen or other related and non-related viral proteins. The inhibition of DNA replication results in a subsequent induction of the host SOS DNA-repair response and in an increase of the mutation frequency in the surviving cells.     

Identification of chalcone synthase genes and their expression patterns reveal pollen abortion in cotton

Chalcone synthase (CHS) is a key enzyme and producing flavonoid derivatives as well play a vital roles in sustaining plant growth and development. However, the systematic and comprehensive analysis of CHS genes in island cotton (G. barbadense) has not been reported yet especially response to cytoplasmic male sterility (CMS). To fill this knowledge gap, a genome-wide investigation of CHS genes were studied in island cotton. A total of 20 GbCHS genes were identified and grouped into five GbCHSs. The gene structure analysis revealed that most of GbCHS genes consisted of two exons and one intron, and 20 motifs were identified. Twenty five pairs duplicated events (12 GbCHS genes) were identified including 23 segmental duplication pairs and two tandem duplication events, representing that GbCHS gene family amplification mainly owned to segmental duplication events and evolving slowly. Gene expression analysis exhibited that the GbCHS family genes presented a diversity expression patterns in various organs of cotton. Coupled with functional predictions and gene expression, the abnormal expression of GbCHS06, 10, 16 and 19 might be associated with pollen abortion of CMS line in island cotton. Conclusively, GbCHS genes exhibited diversity and conservation in many aspects, which will help to better understand functional studies and a reference for CHS research in island cotton and other plants.     

The MDM2 RING Domain and Central Acidic Domain Play Distinct Roles in MDM2 Protein Homodimerization and MDM2-MDMX Protein Heterodimerization

The oncoprotein murine double minute 2 (MDM2) is an E3 ligase that plays a prominent role in p53 suppression by promoting its polyubiquitination and proteasomal degradation. In its active form, MDM2 forms homodimers as well as heterodimers with the homologous protein murine double minute 4 (MDMX), both of which are thought to occur through their respective C-terminal RING (really interesting new gene) domains. In this study, using multiple MDM2 mutants, we show evidence suggesting that MDM2 homo- and heterodimerization occur through distinct mechanisms because MDM2 RING domain mutations that inhibit MDM2 interaction with MDMX do not affect MDM2 interaction with WT MDM2. Intriguingly, deletion of a portion of the MDM2 central acidic domain selectively inhibits interaction with MDM2 while leaving intact the ability of MDM2 to interact with MDMX and to ubiquitinate p53. Further analysis of an MDM2 C-terminal deletion mutant reveals that the C-terminal residues of MDM2 are required for both MDM2 and MDMX interaction. Collectively, our results suggest a model in which MDM2-MDMX heterodimerization requires the extreme C terminus and proper RING domain structure of MDM2, whereas MDM2 homodimerization requires the extreme C terminus and the central acidic domain of MDM2, suggesting that MDM2 homo- and heterodimers utilize distinct MDM2 domains. Our study is the first to report mutations capable of separating MDM2 homo- and heterodimerization.     

The cellular pathway of photosynthate transfer in the developing wheat grain. II. A structural analysis and histochemical studies of the pathway from the crease phloem to the endosperm cavity

In the developing wheat grain, photosynthate is transferred longitudinally along the crease phloem and then laterally into the endosperm cavity through the crease vascular parenchyma, pigment strand and nucellar projection. In order to clarify this cellular pathway of photosynthate unloading, and hence the controlling mechanism of grain filling, the potential for symplastic and apoplastic transfer was examined through structural and histochemical studies on these tissue types. It was found that cells in the crease region from the phloem to the nucellar projection are interconnected by numerous plasmodesmata and have dense cytoplasm with abundant mitochondria. Histochemical studies confirmed that, at the stage of grain development studied, an apoplastic barrier exists in the cell walls of the pigment strand. This barrier is composed of lignin, phenolics and suberin. The potential capacity for symplastic transfer, determined by measuring plasmodesmatal frequencies and computing potential sucrose fluxes through these plasmodesmata, indicated that there is sufficient plasmodesmatal cross-sectional area to support symplastic unloading of photosynthate at the rate required for normal grain growth. The potential capacity for membrane transport of sucrose to the apoplast was assessed by measuring plasma membrane surface areas of the various cell types and computing potential plasma membrane fluxes of sucrose. These fluxes indicated that the combined plasma membrane surface areas of the sieve element–companion cell (se–cc) complexes, vascular parenchyma and pigment strand are not sufficient to allow sucrose transfer to the apoplast at the observed rates. In contrast, the wall ingrowths of the transfer cells in the nucellar projection amplify the membrane surface area up to 22-fold, supporting the observed rates of sucrose transfer into the endosperm cavity. We conclude that photosynthate moves via the symplast from the se–cc complexes to the nucellar projection transfer cells, from where it is transferred across the plasma membrane into the endosperm cavity. The apoplastic barrier in the pigment strand is considered to restrict solute movement to the symplast and block apoplastic solute exchange between maternal and embryonic tissues. The implications of this cellular pathway in relation to the control of photosynthate transfer in the developing grain are discussed.     

LptB-LptF coupling mediates the closure of the substrate-binding cavity in the LptB2FGC transporter through a rigid-body mechanism to extract LPS

Lipopolysaccharides (LPS) are essential envelope components in many Gram-negative bacteria and provide intrinsic resistance to antibiotics. LPS molecules are synthesized in the inner membrane and then transported to the cell surface by the LPS transport (Lpt) machinery. In this system, the ATP-binding cassette (ABC) transporter LptB₂FGC extracts LPS from the inner membrane and places it onto a periplasmic protein bridge through a poorly understood mechanism. Here, we show that residue E86 of LptB is essential for coupling the function of this ATPase to that of its partners LptFG, specifically at the step where ATP binding drives the closure of the LptB dimer and the collapse of the LPS-binding cavity in LptFG that moves LPS to the Lpt periplasmic bridge. We also show that defects caused by changing residue E86 are suppressed by mutations altering either LPS structure or transmembrane helices in LptG. Furthermore, these suppressors also fix defects in the coupling helix of LptF, but not of LptG. Together, these results support a transport mechanism in which the ATP-driven movements of LptB and those of the substrate-binding cavity in LptFG are bi-directionally coordinated through the rigid-body coupling, with LptF’s coupling helix being important in coordinating cavity collapse with LptB dimerization.