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N Xu

Publications and source records attributed to N Xu.

At least 91 records · Page 5Linked to original sources
PubMed1997

Modulation of the fate of cytoplasmic mRNA by AU-rich elements: key sequence features controlling mRNA deadenylation and decay.

Regulation of cytoplasmic deadenylation has a direct impact on the fate of mRNA and, consequently, its expression in the cytoplasm. AU-rich elements (AREs) found in the 3' untranslated regions of many labile mRNAs are the most common RNA-destabilizing elements known in mammalian cells. AREs direct accelerated deadenylation as the first step in mRNA turnover. Recently we have proposed that AREs can be divided into three different classes. mRNAs bearing either the class I AUUUA-containing ARE or the class III non-AUUUA ARE display synchronous poly(A) shortening, whereas class II ARE-containing mRNAs are deadenylated asynchronously, with the formation of poly(A)- intermediates. In this study, we have systematically characterized the deadenylation kinetics displayed by various AREs and their mutant derivatives. We find that a cluster of five or six copies of AUUUA motifs in close proximity forming various degrees of reiteration is the key feature that dictates the choice between processive versus distributive deadenylation. An AU-rich region 20 to 30 nucleotides long immediately 5' to this cluster of AUUUA motifs can greatly enhance the destabilizing ability of the AUUUA cluster and is, therefore, an integral part of the class I and class II AREs. These two features are the defining characteristics of class II AREs. Our results are consistent with the interpretation that the pentanucleotide AUUUA, rather than the nonamer UUAUUUA(U/A)(U/A), is both an essential and the minimal sequence motif of AREs. Our study provides the groundwork for future characterization of ARE-binding proteins identified by in vitro gel shift assays in order to stringently define their potential role in the ARE-mediated decay pathway. Moreover, transformation of deadenylation kinetics from one type to the other by mutations of AREs implies the existence of cross talk between the ARE and 3' poly(A) tail, which dictates the decay kinetics.

3T3 Cells↗
PubMed1997

[Studies on the antimould activity of compound TBZ and its application in paint].

The compound TBZ is a new antimould agent, which contains mainly [2-(4-thiazolyl) benzoimidazole (TBZ)], ethanoic acid and propionic acid. It was shown in laboratory experiment that the compound TBZ had excellent mycostatic effect on all the 10 fungi tested with 70% of the mycostatic zones larger than 6.0 cm. It had obvious inhibitary activity on Alternaria Nees and Aureobasidium Pullulans isolated from mouldy wall. In laboratory condition the surface of fibreboard painted with compound TBZ did not become mouldy in 120 days, while the 20% of that coated with TBZ directly mixed in the paint became mouldy. In practical use, the antimould effect of the wall painted with compound TBZ could maintain more than two years.

Alternaria↗
PubMed1996

The small GTP-binding protein rho activates c-Jun N-terminal kinases/stress-activated protein kinases in human kidney 293T cells. Evidence for a Pak-independent signaling pathway.

Work from a number of laboratories has established a role for certain small GTP-binding proteins in controlling the enzymatic activity of a family of serine-threonine kinases known as mitogen-activated protein kinases (MAPKs). MAPKs have been classified into three subfamilies: extracellular signal-regulated kinases (ERKs), also known as MAPKs; c-Jun N-terminal kinases (JNKs); and p38 kinase. Whereas Ras controls the activation of MAPKs, we and others have recently observed that in certain cells, the small GTP-binding proteins Rac1 and Cdc42 but not Rho regulate the activity of JNKs. Furthermore, because Rac1 and Cdc42 but not Rho bind and activate a kinase known as Pak1, it has been suggested that Pak1 is the most upstream component of the pathway linking these GTPases to JNK. However, in both yeast and mammalian cells, Rho1p, a Rho homologue, and RhoA, respectively, directly interact with a number of proteins, including kinases related to protein kinase C. In addition, in yeast, Rho1p controls the activity of a MAPK cascade involved in bud formation. Considering this diversity of target molecules for small GTP-binding proteins, their likely tissue specific distribution, and the potential role for Rho in signaling to a kinase cascade, we decided to extend our initial analysis, exploring the ability of Ras and Rho-related GTP-binding proteins to activate MAPK or JNK in a variety of cell lines. We found that in the human kidney epithelial cell line, 293T, Cdc42 and all Rho proteins, RhoA, RhoB, and RhoC, but not Rac or Ras can induce activation of JNK. Furthermore, we provide evidence that signaling from Rho proteins to JNK in 293T cells does not involve Pak1. Taken together these findings demonstrate that Rho signals to JNK in a cell type-specific manner and suggest the existence of a novel, Pak1-independent signaling route communicating the Rho family of small GTP-binding proteins to the JNK pathway.

Animals↗
PubMed1996

Trypanothione overproduction and resistance to antimonials and arsenicals in Leishmania.

Leishmania resistant to arsenicals and antimonials extrude arsenite. Previous results of arsenite uptake into plasma membrane-enriched vesicles suggested that the transported species is a thiol adduct of arsenite. In this paper, we demonstrate that promastigotes of arsenite-resistant Leishmania tarentolae have increased levels of intracellular thiols. High-pressure liquid chromatography of the total thiols showed that a single peak of material was elevated almost 40-fold. The major species in this peak was identified by matrix-assisted laser desorption/ionization mass spectrometry as N1,N8-bis-(glutathionyl)spermidine (trypanothione). The trypanothione adduct of arsenite was effectively transported by the As-thiol pump. No difference in pump activity was observed in wild type and mutants. A model for drug resistance is proposed in which Sb(V)/As(V)-containing compounds, including the antileishmanial drug Pentostam, are reduced intracellularly to Sb(III)/As(III), conjugated to trypanothione, and extruded by the As-thiol pump. The rate-limiting step in resistance is proposed to be formation of the metalloid-thiol pump substrates, so that increased synthesis of trypanothione produces resistance. Increased synthesis of the substrate rather than an increase in the number of pump molecules is a novel mechanism for drug resistance.

Animals↗
PubMed1996

Catabolism of prothrombin-chylomicron complexes in vivo in the rat.

The metabolism in vivo in the rat of 125I-prothrombin-chylomicron complexes was compared to that of free 125I-prothrombin injected in saline or together with chyle chylomicrons. The group that obtained 125I-prothrombin-chylomicron complexes exhibited a faster initial decrease of plasma 125I-prothrombin, a higher radioactivity in the liver and more trichloroacetic acid soluble 125I in plasma than in the other groups that did not differ. In the group injected with 125I-prothrombin-chylomicron complexes, 4% was in the HDL, 0.7% LDL, and 0.8% in VLDL 2 hours after injection. The data thus indicate that although most of the 125I-prothrombin is released during the metabolism of the prothrombin-chylomicron complexes, some is rapidly degraded and some is transferred to plasma lipoproteins and remains circulating for a considerable time period. The observations demonstrate that binding to chylomicrons influences the catabolism of prothrombin in vivo. Since chylomicrons did not influence the clearance of simultaneously injected free 125I-prothrombin, the prothrombin binding to chylomicrons or chylomicron remnants in vivo may, however, be too slow to influence the prothrombin catabolism under normal conditions.

Animals↗
PubMed1996

Glutathione and phospholipid depletion of liver tumors after arterial ischemia.

Breakdown of membrane phospholipids is a causative event leading to irreversible cell injury after ischemia and reperfusion insults, which might be one mechanism leading to liver tumor cell death after repeated arterial ischemia as well. After 2 hr of hepatic dearterialization followed by 30 min of reperfusion tumor phospholipid was measured chromatographically, glutathione (GSH) analyzed by determining nonprotein sulfhydryl and activity of glutathione-S-transferase (GST) determined spectrophotometrically using 1-chloro-2,4-dinitrobenzene (CDNB) as the substrate. A transient, arterial ischemia for 2 hr induced a substantial decrease of phosphatidylserine (PS) and phosphatidylinosital (PI) compared with sham treatment (P < 0.01). Although phosphatidylcholine (PC) and phosphatidylethanolamine (PE) did not significantly decline after a single arterial ischemia for 2 hr, they dropped dramatically following repeated arterial ischemia for 2 hr during 5 days (P < 0.01 and P < 0.05 respectively). GSH was depleted in tumors after both a single (P < 0.01) and repeated arterial ischemia (P < 0.05) and GST was inactivated as well (P < 0.001). By contrast, neither liver phospholipid nor liver GSH or GST was significantly changed. Tumor growth was significantly retarded in rats subjected to repeated arterial ischemia compared with sham treatment (P < 0.01). Repeated arterial ischemia facilitated degradation of tumor membrane phospholipids and induced depletion of GSH and inactivation of GST without affecting the normal liver. Thus, ischemia/reperfusion induced depletion of membrane phospholipids and of GSH might represent two mechanisms by which repeated arterial ischemia led to tumor growth delay.

Animals↗
PubMed1996

The PH domain of Ras-GAP is sufficient for in vitro binding to beta gamma subunits of heterotrimeric G proteins.

1. The noncatalytic domain of Ras-GAP can affect signaling through G protein-coupled receptors by a poorly understood mechanism. 2. In this study, fusion proteins containing elements of the noncatalytic domain of ras-GAP were examined for their ability to bind beta gamma subunits of heterotrimeric G proteins and phosphotyrosine-containing polypeptides. 3. Our results demonstrate that purified beta gamma dimers associated with bacterially expressed GAP proteins and that this association does not require SH2 or SH3 domains but is dependent on the presence of the GAP pleckstrin-homology (PH) domain. In contrast, only the SH2 domains are necessary for binding to tyrosine phosphorylated proteins. 4. These findings raise the possibility that heterotrimeric G proteins might affect functioning of ras-like proteins through beta gamma subunits acting on their regulatory molecules.

Binding Sites↗
PubMed1996

Distal cis-acting elements restrict expression of the CyIIIb actin gene in the aboral ectoderm of the sea urchin embryo.

The distal region of the S. purpuratus actin CyIIIb gene, between -400 and -1400 nucleotides, contains at least three distinct cis-acting elements (C1R, C1L and E1) which are necessary for correct expression of fusion reporter genes in transgenic sea urchin embryos. The contribution of these elements in the temporal and spatial regulation of the gene was analyzed by single and double site-directed mutagenesis in fusion constructs which carry the bacterial chloramphenicol acetyl transferase (CAT) gene as a reporter. Following microinjection of the transgenes in sea urchin embryos, the activity of the mutants was compared to the wild type in time and space by measuring CAT activity at the blastula and pluteus embryonic stages and by in situ hybridization to the CAT mRNA at pluteus stage. Our results indicate that E1 is involved in the temporal regulation of CyIIIb and that all three elements are necessary and sufficient to confer aboral (dorsal) ectoderm specificity to the proximal promoter. This is achieved by suppressing the promoter's activity in all other tissues by the cooperative interaction of the cis-acting elements. The C1R element, binding site of the nuclear receptors SpCOUP-TF and SpSHR2, is by itself sufficient to restrict expression in the ectoderm, whereas the aboral ectoderm restricted expression requires in addition the presence of both C1L and E1. It is therefore evident, that the actin CyIIIb gene is exclusively expressed in the aboral ectoderm by a combinatorial repression in all other cell lineages of the developing embryo.

Actins↗
PubMed1996

Endotoxin inhibits catabolism of low density lipoproteins in vivo: an experimental study in the rat.

Hyperlipidaemia frequently accompanies infectious diseases and may be due to an increase in lipoprotein production or a decrease of lipoprotein clearance. The administration of endotoxin has been used to mimic infection and previous studies have demonstrated that endotoxin induces an increase in low density lipoprotein (LDL) levels. In the present study in rats, the dose of endotoxin (055:B5, 50 micrograms 100 g body weight-1) induced hyperlipidaemia without shock and death. The clearance of 125I-LDL was measured after simultaneous injection of 100 micrograms LDL protein with endotoxin or without endotoxin (control). Endotoxin significantly inhibited the clearance of 125I-LDL from blood at all time intervals between 1 and 24 h. The total 125I and trichloroacetic acid-(TCA) precipitable 125I in the liver was higher in endotoxin-treated animals than in controls after 0.5, 4 and 24 h. An increased tissue radioactivity was also seen in several other tissues at various time intervals in the endotoxin treated group. The increase in plasma triglyceride induced by endotoxin reached a maximum after 8 h. Endotoxin thus causes a rapid inhibition of both the LDL disappearance from blood and of the LDL degradation after the uptake by the liver. This effect precedes the increase in triglyceride levels. The inhibition of the LDL catabolism may contribute to the rise in LDL levels in endotoxin induced hyperlipidaemia.

Animals↗
PubMed1996

Effect of exogenous glycine on peptidoglycan composition and resistance in a methicillin-resistant Staphylococcus aureus strain.

A highly homogeneously methicillin-resistant Staphylococcus aureus strain was grown in the presence of various concentrations of exogenous glycine. Increasing concentrations of glycine in the medium resulted in a decrease in methicillin resistance and the appearance of a heterogeneous resistance phenotype. Parallel to the gradual changes in resistance was an alteration in the muropeptide composition of peptidoglycan. Increasing concentrations of glycine in the medium resulted in peptidoglycan in which muropeptides with a D-alanyl-D-alanine terminus were replaced with D-alanyl-glycine-terminating muropeptides. The disappearance of D-alanyl-D-alanine-terminating muropeptides in peptidoglycan and the concomitant decrease in resistance indicate a central role for D-alanyl-D-alanine-terminating precursors in methicillin resistance.

Amino Acid Sequence↗
PubMed1996

[Effects of electroacupuncture at du meridian on contents of nitric oxide and endothelin in rats with acute cerebral ischemia].

Thirty rats were randomly assigned to three groups, control group (10 cases), acute cerebral ischemia group (10 cases) and electroacupuncture (EA) group (10 cases). The bilateral common carotid arteries were occluded, which caused sharp drop of regional cerebral bloodflow and led acute cerebral ischemia. Contents of nitric oxide (NO) and endothalin (ET) in the cerebral cortex and boold were determined in normal, during occluding and after EA of Du meridian (GV 20, GV 14) point in rats. It was found that in acute cerebral ischemia, the brain contents of NO and ET increased, the plasme level of ET elevated and serum level of NO decreased. After EA of Du meridian point, the levels of ET and NO in the brain and blood were returned significantly to normal. It is suggested that EA at Du meridian have protective effect on neural damage induced by brain ischemia. NO and ET are possibly involved in the regulative effect of EA.

Animals↗
PubMed1996

[The effect of phospholipid liposomes on atherosclerosis and serum lipid in rabbits].

In order to study the effect of oral phospholipid liposomes on regression of atherosclerosis process, serum total cholesterol triglyceride, lipoprotein cholesterol and atherosclerotic lesion on the aorta wall in different groups of 20 New Zealand rabbits were observed. Results showed that phospholipid liposomes could obviously increase high density lipoprorein cholesterol concentration and decrease serum total cholesterol triglyceride and very low density lipoprorein cholesterol concentration. Phospholipid liposomes could also reduce the size of atherosclerotic lesion on the aortic wall. In comparision with the results observed in phospholipid group, there was significant difference (P < 0.05, P < 0.01 or P < 0.001). It suggested that oral phospholipid liposomes could effectively regress atherosclerotic lesion on the arterial wall and regulate serum lipid to reduce cholesterol deposit on the arterial wall in experimental rabbits. It might provide some experimental basis for human being to prevent and treat atherosclerosis.

Animals↗
PubMed1995

Dual effect of beta-adrenergic receptors on mitogen-activated protein kinase. Evidence for a beta gamma-dependent activation and a G alpha s-cAMP-mediated inhibition.

The enzymatic activity of mitogen-activated protein kinases (MAP kinases) increases in response to agents acting on a variety of cell surface receptors, including receptors linked to heterotrimeric G proteins of the Gi and Gq family. Recently, it has been shown that stimulation of beta-adrenergic receptors, which are typical of those that act through Gs to activate adenylyl cyclases, potently activates MAP kinases in the heart, resulting in the hypertrophy of the cardiac muscle (Lazou, A., Bogoyevitch, M.A., Clerk, A., Fuller, S.J., Marshall, C.J., and Sudgen, P.H. (1994) Circ. Res. 75, 938-941). We have observed that exposure of COS-7 cells to a beta-adrenergic agonist, isoproterenol, raises intracellular levels of cAMP and effectively activates protein kinase A (PKA) and an epitope-tagged MAP kinase. However, MAP kinase stimulation by isoproterenol was neither mimicked by expression of an activated mutant of G alpha s, nor by treatment with PKA-stimulating agents. Moreover, pretreatment of COS-7 with a permeable cAMP analog, 8-Br-cAMP, markedly decreased MAP kinase activation by either isoproterenol or epidermal growth factor. Thus, in COS-7 cells cAMP and PKA do not appear to mediate MAP kinase activation by beta-adrenergic receptors. Signaling from beta-adrenergic receptors to MAP kinase was inhibited by transfection of a chimeric molecule consisting of the CD8 receptor and the carboxyl terminus of the beta-adrenergic receptor kinase, which includes the beta gamma-binding domain. MAP kinase activation by isoproterenol was not affected by depletion of protein kinase C, but it was completely abolished by expression of Ras-inhibiting molecules. We conclude that signaling from beta-adrenergic receptors to MAP kinase involves an activating signal mediated by beta gamma subunits acting on a Ras-dependent pathway and a G alpha s-induced inhibitory signal mediated by cAMP and PKA. The balance between these two opposing mechanisms of regulation would be expected to control the MAP kinase response to beta-adrenergic agonists as well as to other biologically active agents known to act on Gs coupled receptors, including a number of hormones, neurotransmitters, and lipid mediators.

Base Sequence↗
PubMed1995

Binding of prothrombin to chyle chylomicrons: effects of temperature and calcium ions, and role of surface phospholipids.

The ability of chyle chylomicrons to bind prothrombin has been studied. Rat chyle chylomicrons were incubated with human 125I-prothrombin and binding was examined by separating the chylomicrons from free 125-I-prothrombin by density-gradient ultracentrifugation, and by gel filtration on Sepharose CL-2B. A significant binding of prothrombin to chyle chylomicrons occurred. The complex formation was calcium dependent, and decreased markedly when the temperature was lowered from 37 degrees C to 20 degrees C and when PH was raised above 8. The time course for the binding at 37 degrees C in presence of 2 mmol/L CaCl2 exhibited an initial lag phase at about 10 minutes. Thereafter most of the binding occurred within 30 minutes. Bound prothrombin could not be removed from chyle chylomicrons by treatment with EDTA, suggesting that this binding is not a simple Ca2+ dependent association between prothrombin and chyle chylomicrons. Inclusion of 1% purified human serum albumin caused a 50% decrease in binding, half of which was reversed by increasing the Ca2+ ion concentration. Addition of pancreatic phospholipase A2 (PLA2) in doses sufficient to hydrolyze more than 95% of the phosphatidylethanolamine (PE) and 37% of the phosphatidylcholine (PC) decreased the binding by 50%. Doses of PLA2 that hydrolyze more than 95% of the phosphatidylethanolamine (PE) and 37% of the phosphatidylcholine (PC) decreased the binding by 50%. Doses of PLA2 that hydrolyzed 60-80% of (PE and 4-10% of the PC decreased the binding by only 7-15%. It is suggested that the binding of prothrombin to chyle chylomicrons is in part mediated by negatively charged phospholipids of the chylomicron surface, although a specific role of the PE could not be demonstrated.

Animals↗
PubMed1995

Fatty acid substrate specificities of human prostaglandin-endoperoxide H synthase-1 and -2. Formation of 12-hydroxy-(9Z, 13E/Z, 15Z)- octadecatrienoic acids from alpha-linolenic acid.

Human prostaglandin-endoperoxide H synthase-1 and -2 (hPGHS-1 and hPGHS-2) were expressed by transient transfection of COS-1 cells. Microsomes prepared from the transfected cells were used to measure the rates of oxygenation of several 18- and 20-carbon polyunsaturated fatty acid substrates including eicosapentaenoic, arachidonic, dihomo-gamma-linolenic > alpha-linolenic (delta 9, 12, 15), gamma-linolenic, and linoleic acids. Comparisons of kcat/Km values indicate that the order of efficiency of oxygenation is arachidonate > dihomo-gamma-linolenate > linoleate > alpha-linolenate for both isozymes; while the order of efficiency was the same for hPGHS-1 and hPGHS-2, alpha-linolenate was a particularly poor substrate for hPGHS-1. Gamma-Linolenate and eicosapentaenoate were poor substrates for both isozymes, but in each case, these two fatty acids were better substrates for hPGHS-2 than hPGHS-1. These studies of substrate specificities are consistent with previous studies of the interactions of PGHS isozymes with nonsteroidal anti-inflammatory drugs that have indicated that the cyclooxygenase active site of PGHS-2 is somewhat larger and more accommodating than that of PGHS-1. The major products formed from linoleate and alpha-linolenate were characterized. 13-Hydroxy-(9Z,11E)-octadecadienoic acid was found to be the main product formed from alpha-linoleate by both isozymes. The major products of oxygenation of alpha-linolenate were determined by mass spectrometry to be 12-hydroxy-(9Z,13E/Z,15Z)-octadecatrienoic acids. This result suggests that alpha-linolenate is positioned in the cyclooxygenase active site with a kink in the carbon chain such that hydrogen abstraction occurs from the omega 5-position in contrast to abstraction of the omega 8-hydrogen from other substrates.

Animals↗