Short-chain lipid peroxidation products form covalent adducts with pyruvate kinase and inhibit its activity in vitro and in breast cancer cells

Pyruvate kinase catalyses the last step in glycolysis and has been suggested to contribute to the regulation of aerobic glycolysis in cancer cells. It can be inhibited by oxidation of cysteine residues in vitro and in vivo, which is relevant to the more pro-oxidant state in cancer and proliferating tissues. These conditions also favour lipid peroxidation and the formation of electrophilic fragmentation products, including short-chain aldehydes that can covalently modify proteins. However, as yet few studies have investigated their interactions with pyruvate kinase, so we investigated the effects of three different aldehydes, acrolein, malondialdehyde and 4-hydroxy-2(E)-hexenal (HHE), on the structure and activity of the enzyme. Analysis by LC-MS/MS showed unique modification profiles for each aldehyde, but Cys152, Cys423 and Cys474 were the residues most susceptible to electrophilic modification. Analysis of enzymatic activity under these conditions showed that acrolein was the strongest inhibitor, and at incubation times longer than 2 h, pathophysiological concentrations induced significant effects. Treatment of MCF-7 cells with the aldehydes caused similar losses of pyruvate kinase activity to those observed in vitro, and at lower concentrations than those required to cause cell death, with time and dose-dependent effects; acrolein adducts on Cys152 and Cys358 were detected. Cys358 and Cys474 are located at or near the allosteric or active sites, and formation of adducts on these residues probably contributes to loss of activity at low treatment concentrations. This study provides the first detailed analysis of the structure-activity relationship of C3 and C6 aldehydes with pyruvate kinase, and suggests that reactive short-chain aldehydes generated in diseases with an oxidative aetiology or from environmental exposure such as smoking could be involved in the metabolic alterations observed in cancer cells, through alteration of pyruvate kinase activity

Site 503: Eastern Equatorial Pacific

Our primary objective at Site 503 (Fig. 1) was to re- cover a complete, undisturbed Neogene and Quaternary section in the eastern equatorial Pacific. Site 503 is located near Site 83 in an area that contains an almost continuous pelagic record of the past 10 m.y. (Hays et al., 1972). Unfortunately, Site 83 was only spot-cored, and the recovered sediment is so badly disturbed by rotary drilling that most of the detailed record is lost. The section has an average sedimentation rate of 2.0 to 2.5 cm/k.y. with good-to-moderate preservation of all the major microfossil groups. We returned to Site 83 to core the same section, using the Hydraulic Piston Corer (HPC) to obtain an undisturbed, continuous section for high-resolution stratigraphic studies

Site 502: Colombia Basin, Western Caribbean

Our specific objective at Site 502 was to recover an undisturbed, complete section that could be used as a Neogene and Quaternary reference section. A complete record such as this would allow intercorrelations between (1) paleomagnetic stratigraphy, (2) calcareous biostratigraphy, (3) cyclic accumulation of sediment, (4) paleoceanographic changes, (5) oxygen and carbon isotope stratigraphies, (6) the chronology of Central American volcanism, (7) the timing and effects of the emergence of the Isthmus of Panama, and (8) the timing and effects of the initiation of Northern Hemisphere glaciation

Overexpression of Parathyroid Hormone-related Protein in the Pancreatic Islets of Transgenic Mice Causes Islet Hyperplasia, Hyperinsulinemia, and Hypoglycemia

Parathyroid hormone-related protein (PTHrP) is produced by the pancreatic islet. It also has receptors on islet cells, suggesting that it may serve a paracrine or autocrine role within the islet. We have developed transgenic mice, which overexpress PTHrP in the islet through the use of the rat insulin II promoter (RIP). Glucose homeostasis in these mice is markedly abnormal; RIP-PTHrP mice are hypoglycemic in the postprandial and fasting states and display inappropriate hyperinsulinemia. At the end of a 24-hour fast, blood glucose values are 49 mg/dl in RIP-PTHrP mice, as compared to 77 mg/dl in normal littermates; insulin concentrations at this time are 6.3 and 3.9 ng/ml, respectively. Islet perifusion studies failed to demonstrate abnormalities in insulin secretion. In contrast, quantitative islet histomorphometry demonstrates that the total islet number and total islet mass are 2-fold higher in RIP-PTHrP mice than in their normal littermates. PTHrP very likely plays a normal physiologic role within the pancreatic islet. This role is most likely paracrine or autocrine. PTHrP appears to regulate insulin secretion either directly or indirectly, through developmental or growth effects on islet mass. PTHrP may have a role as an agent that enhances islet mass and/or enhances insulin secretion

Search for CP Violation in the Decay Z -> b (b bar) g

About three million hadronic decays of the Z collected by ALEPH in the years 1991-1994 are used to search for anomalous CP violation beyond the Standard Model in the decay Z -> b \bar{b} g. The study is performed by analyzing angular correlations between the two quarks and the gluon in three-jet events and by measuring the differential two-jet rate. No signal of CP violation is found. For the combinations of anomalous CP violating couplings, ${\hat{h}}_b = {\hat{h}}_{Ab}g_{Vb}-{\hat{h}}_{Vb}g_{Ab}$ and $h^{\ast}_b = \sqrt{\hat{h}_{Vb}^{2}+\hat{h}_{Ab}^{2}}$, limits of \hat{h}_b < 0.59$and$h^{\ast}_{b} < 3.02$ are given at 95\% CL.Comment: 8 pages, 1 postscript figure, uses here.sty, epsfig.st

Activation of Wnt Signaling by Chemically Induced Dimerization of LRP5 Disrupts Cellular Homeostasis

Wnt signaling is crucial for a variety of biological processes, including body axis formation, planar polarity, stem cell maintenance and cellular differentiation. Therefore, targeted manipulation of Wnt signaling in vivo would be extremely useful. By applying chemical inducer of dimerization (CID) technology, we were able to modify the Wnt co-receptor, low-density lipoprotein (LDL)-receptor-related protein 5 (LRP5), to generate the synthetic ligand inducible Wnt switch, iLRP5. We show that iLRP5 oligomerization results in its localization to disheveled-containing punctate structures and sequestration of scaffold protein Axin, leading to robust β-catenin-mediated signaling. Moreover, we identify a novel LRP5 cytoplasmic domain critical for its intracellular localization and casein kinase 1-dependent β-catenin signaling. Finally, by utilizing iLRP5 as a Wnt signaling switch, we generated the Ubiquitous Activator of β-catenin (Ubi-Cat) transgenic mouse line. The Ubi-Cat line allows for nearly ubiquitous expression of iLRP5 under control of the H-2Kb promoter. Activation of iLRP5 in isolated prostate basal epithelial stem cells resulted in expansion of p63+ cells and development of hyperplasia in reconstituted murine prostate grafts. Independently, iLRP5 induction in adult prostate stroma enhanced prostate tissue regeneration. Moreover, induction of iLRP5 in male Ubi-Cat mice resulted in prostate tumor progression over several months from prostate hyperplasia to adenocarcinoma. We also investigated iLRP5 activation in Ubi-Cat-derived mammary cells, observing that prolonged activation results in mammary tumor formation. Thus, in two distinct experimental mouse models, activation of iLRP5 results in disruption of tissue homeostasis, demonstrating the utility of iLRP5 as a novel research tool for determining the outcome of Wnt activation in a precise spatially and temporally determined fashion

The Mechanism for RNA Recognition by ANTAR Regulators of Gene Expression

ANTAR proteins are widespread bacterial regulatory proteins that have RNA–binding output domains and utilize antitermination to control gene expression at the post-initiation level. An ANTAR protein, EutV, regulates the ethanolamine-utilization genes (eut) in Enterococcus faecalis. Using this system, we present genetic and biochemical evidence of a general mechanism of antitermination used by ANTARs, including details of the antiterminator structure. The novel antiterminator structure consists of two small hairpins with highly conserved terminal loop residues, both features being essential for successful antitermination. The ANTAR protein dimerizes and associates with its substrate RNA in response to signal-induced phosphorylation. Furthermore, bioinformatic searches using this conserved antiterminator motif identified many new ANTAR target RNAs in phylogenetically diverse bacterial species, some comprising complex regulons. Despite the unrelatedness of the species in which they are found, the majority of the ANTAR–associated genes are thematically related to nitrogen management. These data suggest that the central tenets for gene regulation by ANTAR antitermination occur widely in nature to specifically control nitrogen metabolism

Measurement of the inclusive isolated-photon cross section in pp collisions at √s = 13 TeV using 36 fb−1 of ATLAS data

The differential cross section for isolated-photon production in pp collisions is measured at a centre-of-mass energy of 13 TeV with the ATLAS detector at the LHC using an integrated luminosity of 36.1 fb. The differential cross section is presented as a function of the photon transverse energy in different regions of photon pseudorapidity. The differential cross section as a function of the absolute value of the photon pseudorapidity is also presented in different regions of photon transverse energy. Next-to-leading-order QCD calculations from Jetphox and Sherpa as well as next-to-next-to-leading-order QCD calculations from Nnlojet are compared with the measurement, using several parameterisations of the proton parton distribution functions. The predictions provide a good description of the data within the experimental and theoretical uncertainties. [Figure not available: see fulltext.