The yeast nuclear cap binding complex can interact with translation factor eIF4G and mediate translation initiation

The mRNA cap structure is bound by either the nuclear (CBC) or the cytoplasmic (eIF4F) cap binding complex. Following mRNA export, CBC must be exchanged for eIF4F in the cytoplasm. It is not known how this exchange occurs or how this RNP remodeling event is integrated with mRNA function. Here we report genetic and biochemical evidence that the yeast translation initiation factor eIF4G associates with CBC, and that eIF4E, the eIF4F component that binds both the cap and eIF4G, antagonizes this interaction. Furthermore, we find that CBC can stimulate translation in extracts containing an eIF4G protein deficient for eIF4E binding. These data suggest that eIF4E binding to the eIF4G-CBC complex on newly exported mRNA displaces CBC, and that the first round of translation on mRNA may occur via a different mechanism than subsequent rounds.     

The immunobiology of mushrooms

There has been enormous interest in the biologic activity of mushrooms and innumerable claims have been made that mushrooms have beneficial effects on immune function with subsequent implications for inhibition of tumor growth. The majority of these observations are anecdotal and often lack standardization. However, there remains considerable data on both in vitro and in vivo effects that reflect on the potential of mushroom compounds to influence human immunity. A number of these effects are beneficial but, unfortunately, many responses are still characterized based on phenomenology and there is more speculation than substance. With respect to tumor biology, although many neoplastic lesions are immunogenic, tumor antigens frequently are self antigens and induce tolerance and many patients with cancer exhibit suppressed immune responses, including defective antigen presentation. Therefore, if and when mushroom extracts are effective, they more likely function as a result of improved antigen presentation by dendritic cells than by a direct cytopathic effect. In this review we attempt to place these data in perspective, with a particular focus on dendritic cell populations and the ability of mushroom extracts to modulate immunity. There is, at present, no scientific basis for the use of either mushrooms or mushroom extracts in the treatment of human patients but there is significant potential for rigorous research to understand the potential of mushrooms in human disease and thence to focus on appropriate clinical trials to demonstrate effectiveness and/ or potential toxicity.     

Studies on the Hemolytic Properties of Protamine

The basic protein protamine causes a rapid hemolysis when incubated with the red blood cells of many mammalian species. The age of the cells does not affect the process. Neutralization of the active side groups of the protamine molecule with formalinization demonstrates that a specific degree of charge is necessary for hemolysis, as more than 30 per cent of the guanidine groups must remain unreacted to maintain activity. Unlike the hemolysis induced by the synthetic polypeptides polylysine and polyhomoarginine, protamine hemolysis is temperature-dependent. Whole lipoprotein material derived from red blood cell membranes inhibits protamine hemolysis to a greater extent than do the membranes themselves, serum, serum protein fractions, or cholesterol. The phosphatide and protein moieties derived from the membranes are quite avid in inhibiting protamine hemolysis. A probable explanation is that intracellular aggregation of these structural elements may cause changes in electrostatic charge and surface tension which result in increased permeability. The hemolytic and antitumor cell properties of protamine could not be segregated from its animal toxicity. Despite formalinization to a degree which eliminated the former, the compound remained quite toxic to mice and rabbits.     

Coupling of Gab1 to c-Met, Grb2, and Shp2 mediates biological responses

Gab1 is a substrate of the receptor tyrosine kinase c-Met and involved in c-Met-specific branching morphogenesis. It associates directly with c-Met via the c-Met-binding domain, which is not related to known phosphotyrosine-binding domains. In addition, Gab1 is engaged in a constitutive complex with the adaptor protein Grb2. We have now mapped the c-Met and Grb2 interaction sites using reverse yeast two-hybrid technology. The c-Met-binding site is localized to a 13-amino acid region unique to Gab1. Insertion of this site into the Gab1-related protein p97/Gab2 was sufficient to confer c-Met-binding activity. Association with Grb2 was mapped to two sites: a classical SH3-binding site (PXXP) and a novel Grb2 SH3 consensus-binding motif (PX(V/I)(D/N)RXXKP). To detect phosphorylation-dependent interactions of Gab1 with downstream substrates, we developed a modified yeast two-hybrid assay and identified PI(3)K, Shc, Shp2, and CRKL as interaction partners of Gab1. In a trk-met-Gab1-specific branching morphogenesis assay, association of Gab1 with Shp2, but not PI(3)K, CRKL, or Shc was essential to induce a biological response in MDCK cells. Overexpression of a Gab1 mutant deficient in Shp2 interaction could also block HGF/SF-induced activation of the MAPK pathway, suggesting that Shp2 is critical for c-Met/Gab1-specific signaling.     

Oxygen-dependent quenching of phosphorescence used to characterize improved myocardial oxygenation resulting from vasculogenic cytokine therapy

This study evaluates a therapy for infarct modulation and acute myocardial rescue and utilizes a novel technique to measure local myocardial oxygenation in vivo. Bone marrow-derived endothelial progenitor cells (EPCs) were targeted to the heart with peri-infarct intramyocardial injection of the potent EPC chemokine stromal cell-derived factor 1α (SDF). Myocardial oxygen pressure was assessed using a noninvasive, real-time optical technique for measuring oxygen pressures within microvasculature based on the oxygen-dependent quenching of the phosphorescence of Oxyphor G3. Myocardial infarction was induced in male Wistar rats (n = 15) through left anterior descending coronary artery ligation. At the time of infarction, animals were randomized into two groups: saline control (n = 8) and treatment with SDF (n = 7). After 48 h, the animals underwent repeat thoracotomy and 20 μl of the phosphor Oxyphor G3 was injected into three areas (peri-infarct myocardium, myocardial scar, and remote left hindlimb muscle). Measurements of the oxygen distribution within the tissue were then made in vivo by applying the end of a light guide to the beating heart. Compared with controls, animals in the SDF group exhibited a significantly decreased percentage of hypoxic (defined as oxygen pressure ≤ 15.0 Torr) peri-infarct myocardium (9.7 ± 6.7% vs. 21.8 ± 11.9%, P = 0.017). The peak oxygen pressures in the peri-infarct region of the animals in the SDF group were significantly higher than the saline controls (39.5 ± 36.7 vs. 9.2 ± 8.6 Torr, P = 0.02). This strategy for targeting EPCs to vulnerable peri-infarct myocardium via the potent chemokine SDF-1α significantly decreased the degree of hypoxia in peri-infarct myocardium as measured in vivo by phosphorescence quenching. This effect could potentially mitigate the vicious cycle of myocyte death, myocardial fibrosis, progressive ventricular dilatation, and eventual heart failure seen after acute myocardial infarction.     

Female genital system of the folding-trapdoor spider Antrodiaetus unicolor (Hentz, 1842) (Antrodiaetidae, Araneae): ultrastructural study of form and function with notes on reproductive biology of spiders

The genitalia of the female folding-trapdoor spider Antrodiaetus unicolor are characterized by two pairs of spermathecae that are arranged in a single row and connected to the roof of the bursa copulatrix. Each single spermatheca is divided into three main parts: stalk, bowl, and bulb, which are surrounded by the spermathecal gland. The epithelium of the spermathecal gland is underlain by a muscle meshwork and consists of different types of cells partly belonging to glandular cell units (Class 3 gland cells) that extend into pores in the cuticle of the stalk and bowl. Interestingly, the bulb lacks glandular pores and is characterized by a weakly sclerotized cuticle. This peculiarly structured bulb probably plays an important role in the discharge of the sperm mass. It is suggested that by contraction of the muscle layer the sperm mass may be squeezed out, when the bulb invaginates and expands into the spermathecal lumen, pushing the sperm to the uterus lumen. Each glandular unit consists of usually one or two central secretory cells that are for the most part surrounded by a connecting cell that again is surrounded by a canal cell. The canal cell, finally, is separated from the other epithelial cells (intercalary cells) located between the glandular units by several thin sheath cells that form the outer enveloping layer of the unit. The secretions are released through a cuticular duct that originates proximally between the apical part of the connecting cell and the apical microvilli of the secretory cells and runs into a pore of the spermathecal cuticle. The glandular products of the Class 3 gland cells likely contribute to the conditions allowing long-term storage of the spermatozoa in this species. Details regarding the ovary, the uterus internus, and the uterus externus are reported. Most of the secretion that composes the chorion of the egg is produced in the ovary. Glandular cell units observed in the uterus externus differ structurally from those in the spermathecae and likely play a different role. Finally, we briefly discuss our results on the female genitalia of A. unicolor in the light of knowledge about the reproductive biology of spiders.     

The taxonomic and ecological status of the environmentally restricted spongillid species of North America. IV. Spongilla heterosclerifera Smith 1918

The taxonomic status, present distribution and specific threats to the existence of the environmentally restricted freshwater sponge, Spongilla heterosclerifera Smith 1918 were investigated. This species, collected only from Oneida Lake, New York, has not colonized other habitats and continues to exhibit typical diagnostic characteristics, thus qualifying as a valid environmentally restricted species. Although the sponge presently colonizes two sites in the lake, both near the northwestern shore, the total absence of sponge fauna from other lake regions near more heavily populated areas of this species. Physicochemical data from Oneida Lake greatly extend the known environmental parameters of Spongilla heterosclerifera.     

p75 neurotrophin receptor regulates agonist-induced pulmonary vasoconstriction

A member of the TNF receptor family, the p75 neurotrophin receptor (p75(NTR)) has been previously shown to play a role in the regulation of fibrin deposition in the lung. However, the role of p75(NTR) in the regulation of pulmonary vascular tone in the lung is unknown. In the present study, we evaluated the expression of p75(NTR) in mouse pulmonary arteries and the putative role of p75(NTR) in modulating pulmonary vascular tone and agonist responsiveness using wild-type (WT) and p75(NTR) knockout (p75(-/-)) mice. Our data indicated that p75(NTR) is expressed in both smooth muscle and endothelial cells within the pulmonary vascular wall in WT mice. Pulmonary artery rings from p75(-/-) mice exhibited significantly elevated active tension due to endothelin-1-mediated Ca(2+) influx. Furthermore, the contraction due to capacitative Ca(2+) entry (CCE) in response to phenylephrine-mediated active depletion of intracellular Ca(2+) stores was significantly enhanced compared with WT rings. The contraction due to CCE induced by passive store depletion, however, was comparable between WT and p75(-/-) rings. Active tension induced by serotonin, U-46619 (a thromboxane A(2) analog), thrombin, 4-aminopyridine (a K(+) channel blocker), and high extracellular K(+) in p75(-/-) rings was similar to that in WT rings. Deletion of p75(NTR) did not alter pulmonary vasodilation to sodium nitroprusside (a nitric oxide donor). These data suggest that intact p75(NTR) signaling may play a role in modulating pulmonary vasoconstriction induced by endothelin-1 and by active store depletion.