Syk-dependent Phosphorylation of CLEC-2: A Novel Mechanism of Hem-Immunoreceptor Tyrosine-Based Activation Motif Signaling

The C-type lectin-like receptor CLEC-2 signals via phosphorylation of a single cytoplasmic YXXL sequence known as a hem-immunoreceptor tyrosine-based activation motif (hemITAM). In this study, we show that phosphorylation of CLEC-2 by the snake toxin rhodocytin is abolished in the absence of the tyrosine kinase Syk but is not altered in the absence of the major platelet Src family kinases, Fyn, Lyn, and Src, or the tyrosine phosphatase CD148, which regulates the basal activity of Src family kinases. Further, phosphorylation of CLEC-2 by rhodocytin is not altered in the presence of the Src family kinase inhibitor PP2, even though PLCγ2 phosphorylation and platelet activation are abolished. A similar dependence of phosphorylation of CLEC-2 on Syk is also seen in response to stimulation by an IgG mAb to CLEC-2, although interestingly CLEC-2 phosphorylation is also reduced in the absence of Lyn. These results provide the first definitive evidence that Syk mediates phosphorylation of the CLEC-2 hemITAM receptor with Src family kinases playing a critical role further downstream through the regulation of Syk and other effector proteins, providing a new paradigm in signaling by YXXL-containing receptors

Potential role of endocrine gastrin in the colonic adenoma carcinoma sequence

The role of hyper-gastrinaemia in the incidence of colonic cancer remains to be clarified. The aim of this study was to determine whether cholecystokinin-2 (CCK-2) receptor expression predicts the sensitivity of human colonic adenomas to the proliferative effects of serum hyper-gastrinaemia. Gene expression of the classical (74 kDa) CCK-2 receptor in human colonic adenoma specimens and cell lines, was quantified by real-time PCR. Western blotting, using a CCK-2 receptor antiserum, confirmed protein expression. A transformed human colonic adenoma was grown in SCID mice, with hyper-gastrinaemia induced by protein pump inhibitors. CCK-2 receptor blockade was achieved by using neutralising antiserum. Both human colonic adenoma cell lines and biopsies expressed CCK-2 receptor mRNA at levels comparable with CCK-2 receptor transfected fibroblasts and oxyntic mucosa. Western blotting confirmed immunoreactive CCK-2 receptor bands localised to 45, 74 and 82.5 kDa. Omeprazole and lansoprazole-induced hyper-gastrinaemia (resulting in serum gastrin levels of 34.0 and 153.0 pM, respectively) significantly increased the weight of the human adenoma grafts (43% (P=0.016) and 70% (P=0.014), respectively). The effect of hypergastrinaemia on tumour growth was reversed by use of antiserum directed against the CCK-2 receptor. Hyper-gastrinaemia may promote proliferation of human colonic adenomas that express CCK-2 receptor isoforms

Textures And Traction: How Tube-Dwelling Polychaetes Get A Leg Up

By controlling the traction between its body and the tube wall, a tube-dwelling polychaete can move efficiently from one end of its tube to the other, brace its body during normal functions (e.g., ventilation and feeding), and anchor within its tube avoiding removal by predators. To examine the potential physical interaction between worms and the tubes they live in, scanning electron microscopy was used to reveal and quantify the morphology of worm bodies and the tubes they produce for species representing 13 families of tube-dwelling polychaetes. In the tubes of most species there were macroscopic or nearly macroscopic (~10 μm–1 mm) bumps or ridges that protruded slightly into the lumen of the tube; these could provide purchase as a worm moves or anchors. At this scale (~10 μm-1 mm), the surfaces of the chaetal heads that interact with the tube wall were typically small enough to fit within spaces between these bumps (created by the inward projection of exogenous materials incorporated into the tube wall) or ridges (made by secretions on the interior surface of the tube). At a finer scale (0.01–10 μm), there was a second overlap in size, usually between the dentition on the surfaces of chaetae that interact with the tube walls and the texture provided by the secreted strands or microscopic inclusions of the inner linings. These linings had a surprising diversity of micro-textures. The most common micro-texture was a “fabric” of secreted threads, but there were also orderly micro-ridges, wrinkles, and rugose surfaces provided by microorganisms incorporated into the inner tube lining. Understanding the fine structures of tubes in conjunction with the morphologies of the worms that build them gives insight into how tubes are constructed and how worms live within them

COX-2 selective inhibition reverses the trophic properties of gastrin in colorectal cancer

Gastrin is a gastrointestinal peptide that possesses potent trophic properties on both normal and neoplastic cells of gastrointestinal origin. Previous studies have indicated that chronic hypergastrinaemia increases the risk of colorectal cancer and cancer growth and that interruption of the effects of gastrin could be a potential target in the treatment of colorectal cancer. Here we demonstrate that gastrin leads to a dose-dependent increase in colon cancer cell proliferation and tumour growth in vitro and in vivo, and that this increment is progressively reversed by pretreatment with the cyclo-oxygenase-2 inhibitor NS-398. Gastrin was able to induce cyclo-oxygenase-2 protein expression, as well as the synthesis of prostaglandin E2, the major product of cyclo-oxygenase. Moreover, gastrin leads to approximately a two-fold induction of cyclo-oxygenase-2 promoter activity in transiently transfected cells. The results of these studies demonstrate that cyclo-oxygenase-2 appears to represent one of the downstream targets of gastrin and that selective cyclo-oxygenase-2 inhibition is capable of reversing the trophic properties of gastrin and presumably might prevent the growth of colorectal cancer induced by hypergastrinaemia

Functions of cytochrome c oxidase assembly factors

Cytochrome c oxidase is the terminal complex of eukaryotic oxidative phosphorylation in mitochondria. This process couples the reduction of electron carriers during metabolism to the reduction of molecular oxygen to water and translocation of protons from the internal mitochondrial matrix to the inter-membrane space. The electrochemical gradient formed is used to generate chemical energy in the form of adenosine triphosphate to power vital cellular processes. Cytochrome c oxidase and most oxidative phosphorylation complexes are the product of the nuclear and mitochondrial genomes. This poses a series of topological and temporal steps that must be completed to ensure efficient assembly of the functional enzyme. Many assembly factors have evolved to perform these steps for insertion of protein into the inner mitochondrial membrane, maturation of the polypeptide, incorporation of co-factors and prosthetic groups and to regulate this process. Much of the information about each of these assembly factors has been gleaned from use of the single cell eukaryote Saccharomyces cerevisiae and also mutations responsible for human disease. This review will focus on the assembly factors of cytochrome c oxidase to highlight some of the outstanding questions in the assembly of this vital enzyme complex

Nucleosomes in serum as a marker for cell death

The concentration of nucleosomes is elevated in blood of patients with diseases which are associated with enhanced cell death. In order to detect these circulating nucleosomes, we used the Cell Death Detection-ELISA(Plus) (CDDE) from Roche Diagnostics (Mannheim, Germany) (details at http:\textbackslash{}\textbackslash{}biochem.roche.com). For its application in liquid materials we performed various modifications: we introduced a standard curve with nucleosome-rich material, which enabled direct quantification and improved comparability of the values within (CVinterassay:3.0-4.1%) and between several runs (CVinterassay:8.6-13.5%), and tested the analytical specificity of the ELISA. Because of the fast elimination of nucleosomes from circulation and their limited stability, we compared plasma and serum matrix and investigated in detail the pre-analytical handling of serum samples which can considerably influence the test results. Careless venipuncture producing hemolysis, delayed centrifugation and bacterial contamination of the blood samples led to false-positive results; delayed stabilization with EDTA and insufficient storage conditions resulted in false-negative values. At temperatures of -20 degreesC, serum samples which were treated with 10 mM EDTA were stable for at least 6 months. In order to avoid possible interfering factors, we recommend a schedule for the pre-analytical handling of the samples. As the first stage, the possible clinical application was investigated in the sera of 310 persons. Patients with solid tumors (n = 220; mean = 361 Arbitrary Units (AU)) had considerably higher values than healthy persons (n = 50; mean = 30 AU; P = 0.0001) and patients with inflammatory diseases (n = 40; mean = 296 AU; p = 0.096). Within the group of patients with tumors, those in advanced stages (UICC 4) showed significantly higher values than those in early stages (UICC 1-3) (P = 0.0004)

Maternal and early life nutrition and physical activity: setting the research and intervention agenda for addressing the double burden of malnutrition in South African children.

Early life is important for later health outcomes, yet there are few studies which adequately address all of the potential early life insults that may affect later life health and growth trajectories. This is particularly evident in low- to middle-income countries such as South Africa, where women of childbearing age are particularly vulnerable to high levels of physical inactivity, malnutrition, and obesity. Pregnancy may therefore be an opportune time to change behaviours and improve maternal and offspring health outcomes, and decrease the inter-generational transfer of risk. We show clear evidence that physical activity and nutrition are important target areas for intervention during pregnancy and in the early years of life, yet that current literature in Africa, and specifically South Africa, is limited. We have outlined the available literature concerning the impact of maternal and early life nutrition and physical activity on the health status of South African children, and have provided some recommendations for future research and policy

Ocean acidification boosts larval fish development but reduces the window of opportunity for successful settlement.

Author version of article published in final form at DOI: 10.1098/rspb.2015.1954© 2015 The Author(s) Published by the Royal Society. All rights reservedLocating appropriate settlement habitat is a crucial step in the life cycle of most benthic marine animals. In marine fish, this step involves the use of multiple senses, including audition, olfaction and vision. To date, most investigations of larval fish audition focus on the hearing thresholds to various frequencies of sounds without testing an ecological response to such sounds. Identifying responses to biologically relevant sounds at the development stage in which orientation is most relevant is fundamental. We tested for the existence of ontogenetic windows of reception to sounds that could act as orientation cues with a focus on vulnerability to alteration by human impacts. Here we show that larvae of a catadromous fish species (barramundi, Lates calcarifer) were attracted towards sounds from settlement habitat during a surprisingly short ontogenetic window of approximately 3 days. Yet, this auditory preference was reversed in larvae reared under end-of-century levels of elevated CO2, such that larvae are repelled from cues of settlement habitat. These future conditions also reduced the swimming speeds and heightened the anxiety levels of barramundi. Unexpectedly, an acceleration of development and onset of metamorphosis caused by elevated CO2 were not accompanied by the earlier onset of attraction towards habitat sounds. This mismatch between ontogenetic development and the timing of orientation behaviour may reduce the ability of larvae to locate habitat or lead to settlement in unsuitable habitats. The misinterpretation of key orientation cues can have implications for population replenishment, which are only exacerbated when ontogenetic development decouples from the specific behaviours required for location of settlement habitats.Australian Research Counci