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Biomedical subjects

X Lu

Publications and source records attributed to X Lu.

At least 19 recordsLinked to original sources
PubMed2001

AsMamDB: an alternative splice database of mammals.

The objective of database AsMamDB is to facilitate the systematic study of alternatively spliced genes of mammals. Version 1.0 of AsMamDB contains 1563 alternatively spliced genes of human, mouse and rat, each associated with a cluster of nucleotide sequences. The main information provided by AsMamDB includes gene alternative splicing patterns, gene structures, locations in chromosomes, products of genes and tissues where they express. Alternative splicing patterns are represented by multiple alignments of various gene transcripts and by graphs of their topological structures. Gene structures are illustrated by exon, intron and various regulatory elements distributions. There are 4204 DNAs, 3977 mRNAs, 8989 CDSs and 126 931 ESTs in the current database. More than 130 000 GenBank entries are covered and 4443 MEDLINE records are linked. DNA, mRNA, exon, intron and relevant regulatory element sequences are provided in FASTA format. More information can be obtained by using the web-based multiple alignment tool Asalign and various category lists. AsMamDB can be accessed at http://166.111.30.65/ASMAMDB.html.

Alternative Splicing↗
PubMed2000

Mdm2 inhibits the apoptotic function of p53 mainly by targeting it for degradation.

The ability of Mdm2 to inhibit the activities of a C-terminal truncated p53 mutant, p53-Delta30, which can bind Mdm2 but is resistant to Mdm2-mediated protein degradation was investigated. The inhibitory function of an Mdm2 mutant, Mdm2-Delta(222-437), which can bind p53 but is defective in targeting p53 for degradation was also studied. We have demonstrated that targeting p53 for degradation is the most effective way for Mdm2 to inhibit the apoptotic function of p53. However, we have also shown that Mdm2 can inhibit the transactivation function of p53 without targeting it for degradation, although Mdm2 releases the transrepression ability of p53 mainly by targeting it for degradation. The ability of Mdm2 to inhibit the apoptotic function of p53 was linked to its ability to inhibit the transrepression but not the transactivation function of p53. Furthermore, we have demonstrated that the transrepression function of p53 was specific to p53-induced apoptosis and was not simply a result of cell death.

Animals↗
PubMed2000

Study on the fluorescence spectra and electrochemical behavior of ZnL2 and Morin with DNA.

The interactions of Morin (2', 3, 4', 5, 7-pentahydroxyflavone) and its Zn-complex, ZnL2.3H2O [L = Morin (2'-OH group deprotonated)], with calf thymus DNA have been studied using fluorimetric and electrochemical methods. ZnL2.3H2O has different spectral characteristics and electrochemical behavior from that of Morin in the presence of DNA. Increasing fluorescence is seen for ZnL2.3H2O with DNA addition whereas decreased fluorescence is observed for Morin. An isosbestic point appears at 560 nm for the DNA-ZnL2.3H2O system with ethidium bromide (EB) addition while no isosbestic point is detectable for Morin. Quenching fluorescence is observed for DNA-EB system when ZnL2.3H2O is added whereas no quenched fluorescence is seen for DNA-EB system with Morin addition. The above results suggest that Morin and ZnL2.3H2O can both bind to DNA, but the binding mode is different. The complex binds to DNA mainly by intercalation, while Morin binds in a non-intercalating mode. The binding constant and binding site sizes are derived from electrochemical methods. For ZnL2.3H2O, the binding constant is 5.0 x 10(4) l mol(-1) at 20 degrees C, and the binding site size n(s) is 5.

DNA↗
PubMed2000

Speciation of key arsenic metabolic intermediates in human urine.

Biomethylation is the major human metabolic pathway for inorganic arsenic, and the speciation of arsenic metabolites is essential to a better understanding of arsenic metabolism and health effects. Here we describe a technique for the speciation of arsenic in human urine and demonstrate its application to the discovery of key arsenic metabolic intermediates, monomethylarsonous acid (MMAIII) and dimethylarsinous acid (DMAIII), in human urine. The study provides a direct evidence in support of the proposed arsenic methylation pathway in the human. The finding of MMAIII and DMAIII in human urine, along with recent studies showing the high toxicity of these arsenicals, suggests that the usual belief of arsenic detoxification by methylation needs to be reconsidered. The arsenic speciation technique is based on ion pair chromatographic separation of arsenic species on a 3-micron particle size column at 50 degrees C followed by hydride generation atomic fluorescence detection. Speciation of MMAIII, DMAIII, arsenite (AsIII), arsenate (AsV), monomethylarsonic acid (MMAV), and dimethylarsinic acid (DMAV) in urine samples is complete in 6 min with detection limits of 0.5-2 micrograms/L. There is no need for any sample pretreatment. The capability of rapid analysis of trace levels of arsenic species, which resulted in the findings of the key metabolic intermediates, makes the technique useful for routine arsenic speciation analysis required for toxicological and epidemiological studies.

Arsenic↗
PubMed2000

Ecotropic murine leukemia virus receptor is physically associated with caveolin and membrane rafts.

We used a Sindbis virus expression system to stably express a chimeric ecotropic murine leukemia virus (MLV) receptor gene, CAT1, fused to green fluorescent protein (gfp) in BHK cells. The chimeric gene was expressed on the cell surface and functioned as an MLV receptor. Using gfp as an epitope tag, we found that CAT1 cross-immunoprecipitated with caveolin, a cellular protein associated non-clathrin-coated endocytic vesicles. Biochemical studies showed that CAT1 copurified with caveolin in a detergent-insoluble membrane fraction that forms cholesterol-rich "rafts" on the cell surface. Disruption of rafts by methyl-beta-cyclodextrin, a drug that extracts cholesterol, reduced susceptibility to MLV without decreasing surface CAT1. The results indicate that association of the MLV receptor with cholesterol-rich rafts is important for an early step in virus infection.

Animals↗
PubMed2000

Palladium(II)-catalyzed tandem intramolecular aminopalladation of alkynes and conjugate addition. Synthesis of oxazolidinones, imidazolidinones, and lactams

[reaction: see text]Under the catalysis of a divalent palladium species, oxazolidinones, imidazolidinones, or lactams were conveniently obtained with high chemo- and stereoselectivity from the tandem intramolecular aminopalladation of alkynes, followed by insertion of alkenes, and protonolysis of the newly formed carbon-palladium bond.

Journal Article↗
PubMed2000

Structure-based design of an osteoclast-selective, nonpeptide src homology 2 inhibitor with in vivo antiresorptive activity.

Targeted disruption of the pp60(src) (Src) gene has implicated this tyrosine kinase in osteoclast-mediated bone resorption and as a therapeutic target for the treatment of osteoporosis and other bone-related diseases. Herein we describe the discovery of a nonpeptide inhibitor (AP22408) of Src that demonstrates in vivo antiresorptive activity. Based on a cocrystal structure of the noncatalytic Src homology 2 (SH2) domain of Src complexed with citrate [in the phosphotyrosine (pTyr) binding pocket], we designed 3',4'-diphosphonophenylalanine (Dpp) as a pTyr mimic. In addition to its design to bind Src SH2, the Dpp moiety exhibits bone-targeting properties that confer osteoclast selectivity, hence minimizing possible undesired effects on other cells that have Src-dependent activities. The chemical structure AP22408 also illustrates a bicyclic template to replace the post-pTyr sequence of cognate Src SH2 phosphopeptides such as Ac-pTyr-Glu-Glu-Ile (1). An x-ray structure of AP22408 complexed with Lck (S164C) SH2 confirmed molecular interactions of both the Dpp and bicyclic template of AP22408 as predicted from molecular modeling. Relative to the cognate phosphopeptide, AP22408 exhibits significantly increased Src SH2 binding affinity (IC(50) = 0.30 microM for AP22408 and 5.5 microM for 1). Furthermore, AP22408 inhibits rabbit osteoclast-mediated resorption of dentine in a cellular assay, exhibits bone-targeting properties based on a hydroxyapatite adsorption assay, and demonstrates in vivo antiresorptive activity in a parathyroid hormone-induced rat model.

Adsorption↗
PubMed2000

A facile highly regio- and stereoselective preparation of N-tosyl allylic amines from allylic alcohols and tosyl isocyanate via Palladium(II)-catalyzed aminopalladation-beta-heteroatom elimination

The high regio- and stereoselectivity have been obtained from the allylic substitution reaction catalyzed by palladium(II) species. From allylic alcohols, one-pot reaction with tosyl isocyanate followed by palladium(II)-catalyzed allylic substitution gives N-tosyl (E)-allylic amines in high yield. The substitution occurs only at the gamma-position of the 1- or 3-substituted allylic alcohols.

Journal Article↗
PubMed2000

TRH regulates Kv1.5 gene expression through a Galphaq-mediated PLC-independent pathway.

Thyrotropin-releasing hormone (TRH) decreases transcription of the Kv1.5 K(+) channel gene in GH(3) pituitary cells. Here, we examine whether TRH utilizes Gq activated phospholipase C, Gs or Gi to produce this response. We report that expression of constitutively active Galphaq mimicked and occluded the TRH effect. In contrast, expression of activated Galpha(S) or Galpha(i2) had no effect on Kv1. 5 mRNA expression. Furthermore, pertussis and cholera toxins failed to block the TRH-induced decrease in channel mRNA. Surprisingly, despite the role of Gq, the phospholipase C inhibitor U73122 did not alter down-regulation of channel mRNA by TRH, although it abolished the TRH-induced increase in intracellular [Ca(2+)] and up-regulation of c-fos mRNA. Furthermore, depletion of an intracellular Ca(2+) pool or inhibition of protein kinase C did not block the TRH-induced decrease in Kv1.5 mRNA. These results indicate that TRH-induced down-regulation of Kv1.5 gene expression is mediated by Galphaq proteins, but does not require PLC activation.

Animals↗
PubMed2000

Calpain-mediated degradation of PSD-95 in developing and adult rat brain.

PSD-95 is a major postsynaptic density protein that is degraded as a result of synaptic activity. We used four different methods to test the hypothesis that calpain is involved in PSD-95 turnover. Treatment of synaptic membranes with purified calpain resulted in a decrease in immunoreactivity of the native 95 kDa protein and the appearance of two smaller molecular weight species, migrating at 50 and 36 kDa, respectively. Calcium treatment of frozen-thawed brain sections produced an identical digestion pattern, an effect blocked by calpain inhibitors. N-methyl-D-aspartate treatment of organotypic hippocampal cultures produced truncation of PSD-95 and accumulation of the 36 kDa species. Finally, calpain-generated degradation products of PSD95 were prominent in neonatal hippocampus, and disappeared with postnatal development. Our data suggest that PSD-95 is a substrate for calpain, and that calpain-mediated truncation contributes to PSD-95 turnover.

Age Factors↗
PubMed2000

Cloning of human Ca2+ homoeostasis endoplasmic reticulum protein (CHERP): regulated expression of antisense cDNA depletes CHERP, inhibits intracellular Ca2+ mobilization and decreases cell proliferation.

A monoclonal antibody which blocks InsP(3)-induced Ca(2+) release from isolated endoplasmic reticulum was used to isolate a novel 4.0 kb cDNA from a human erythroleukaemia (HEL) cell cDNA expression library. A corresponding mRNA transcript of approx. 4.2 kb was present in all human cell lines and tissues examined, but cardiac and skeletal muscle had an additional transcript of 6.4 kb. The identification in GenBank(R) of homologous expressed sequence tags from many tissues and organisms suggests that the gene is ubiquitously expressed in higher eukaryotes. The gene was mapped to human chromosome 19p13.1. The cDNA predicts a 100 kDa protein, designated Ca(2+) homoeostasis endoplasmic reticulum protein (CHERP), with two putative transmembrane domains, multiple consensus phosphorylation sites, a polyglutamine tract of 12 repeats and regions of imperfect tryptophan and histadine octa- and nona-peptide repeats. In vitro translation of the full-length cDNA produced proteins of M(r) 128000 and 100000, corresponding to protein bands detected by Western blotting of many cell types. CHERP was co-localized in HEL cells with the InsP(3) receptor by two-colour immunofluorescence. Transfection of HEL cells with antisense cDNA led to an 80% decline in CHERP within 5 days of antisense induction, with markedly decreased intracellular Ca(2+) mobilization by thrombin, decreased DNA synthesis and growth arrest, indicating that the protein has an important function in Ca(2+) homoeostasis, growth and proliferation.

Amino Acid Sequence↗
PubMed2000

Stress signals induce transcriptionally inactive E2F-1 independently of p53 and Rb.

One of the common features of cellular response to stress is cell cycle arrest or apoptosis. E2F is one of the key factors which controls cell cycle progression. Overexpression of E2F-1 can also induce apoptosis. In order to understand the role of E2F-1 in cellular response to stress, we studied the E2F-1 response in various cell lines to different types of stress signals including UV irradiation, cisplatin, etoposide and hypoxia. We showed here that the expression level of E2F-1 can be up regulated by the treatment of DNA damage agents as well as hypoxia. The kinetics of E2F-1 increase was dependent on the types of inducer and was similar to that of p53. However, stress signals can induce E2F-1 expression independently of p53 and Rb. Furthermore, the induced E2F-1 was transcriptionally inactive. All these results suggested that E2F-1 may play a very important role in cellular response to stress and this novel role of E2F-1 is independent of its transactivation function.

Carrier Proteins↗
PubMed2000

Calpain-mediated truncation of rat brain AMPA receptors increases their Triton X-100 solubility.

Previous studies have indicated that calpain activation results in the truncation of the C-terminal domains of AMPA and NMDA receptor subunits. The present study determined the distribution of the truncated species of the subunits between Triton-soluble and -insoluble fractions. Western blots were performed with various antibodies to quantify the amounts of the various species of GluR1, GluR2, GluR3 and NR2B subunits. The results indicate that calpain activation decreased the amount of all the intact subunits in Triton-insoluble fractions. Calpain-generated truncated forms of GluR1 and GluR2, but not NR2B, were absent in these fractions, and were recovered in Triton-soluble fractions. These findings suggest that calpain-mediated truncation of AMPA but not NMDA receptor C-terminal domains results in modifications of the interactions between the receptors and postsynaptic densities, and that this mechanism could be involved in activity-dependent changes in the subcellular distribution of AMPA receptors.

Animals↗
PubMed2000

A nonionic block co-polymer adjuvant (CRL1005) enhances the immunogenicity and protective efficacy of inactivated influenza vaccine in young and aged mice.

The use of adjuvants is one approach to improve influenza vaccine immunogenicity and efficacy, particularly in aged populations. The response of BALB/c mice to subcutaneously administered formalin-inactivated whole influenza virus vaccine in the presence or absence of a nonionic block copolymer adjuvant CRL1005 was evaluated. In young adult naïve mice, the copolymer adjuvant significantly enhanced virus-specific IgG and hemagglutination-inhibition (HI) antibody responses and augmented the production of IL-2 following vaccination. Influenza vaccine formulated with 2.5 mg CRL1005 significantly enhanced the protective efficacy of the inactivated vaccine in the upper and lower respiratory tract. In mice previously infected with influenza virus or naïve aged mice, inactivated vaccine administered with the copolymer adjuvant substantially enhanced the serum HI antibody response to inactivated influenza vaccine and significantly reduced lung virus titers following subsequent challenge with live virus compared with mice administered vaccine alone. These results suggest that the copolymer adjuvant warrants further investigation as a potential adjuvant for use in human vaccination against influenza.

Adjuvants, Immunologic↗
PubMed2000

Phosphorescent oxygen sensors utilizing sulfur-nitrogen-phosphorous polymer matrixes: synthesis, characterization, and evaluation of poly(thionylphosphazene)-b-poly(tetrahydrofuran) block copolymers

We examine the use of thionylphosphazene-based block copolymers as matrixes for oxygen sensor applications. Poly(aminothionylphosphazene)-b-poly(tetrahydrofuran) (PATPy-PTHFx) block copolymers were prepared via reaction of ring-opened poly(chlorothionylphosphazene) with THF and subsequently with excess n-butylamine (to form PBATPy-PTHFx) or methylamine (to form PMATPy-PTHFx). The block copolymers were characterized by NMR, gel permeation chromatography, and differential scanning calorimetry. Films of PBATPy-PTHFx block copolymers containing platinum octaethylporphyrin or [Ru(dpp)3]Cl2 (dpp = 4,7-diphenyl-1,10-phenanthroline) as the oxygen-sensitive chromophore were prepared, and time-scan experiments were carried out to determine the diffusion coefficients, Do2, and solubilities, So2, of oxygen therein. Despite microphase separation, the data fit well to a simple Fick's law description of oxygen diffusion and gave Do2 values smaller than that for the n-butylamino-substituted PBATP635. For films freshly annealed above the melting point of PTHFx, the Do2 values were 35-50% (dye-dependent) larger than after aging 3 days at room temperature. Films with [Ru(dpp)3]Cl2 as the dye were evaluated as media for phosphorescent pressure-sensing. The dye-containing polymer films exhibit linear Stern-Volmer-like plots, even at high dye concentrations, as well as good photostability, and significantly higher sensitivity to oxygen quenching than simple mixtures of the analogous homopolymers.

Journal Article↗
PubMed2000

An artificial transcriptional activating region with unusual properties.

We describe a series of transcriptional activators generated by adding amino acids (eight in one case, six in another) to fragments of the yeast Saccharomyces cerevisiae activator Gal4 that dimerize and bind DNA. One of the novel activating regions identified by this procedure is unusual, compared with previously characterized yeast activating regions, in the following ways: it works more strongly than does Gal4's natural activating region as assayed in yeast; it is devoid of acidic residues; and several lines of evidence suggest that it sees targets in the yeast transcriptional machinery at least partially distinct from those seen by Gal4's activating region.

Base Sequence↗