Biomedical subjects

Y Geng

Publications and source records attributed to Y Geng.

At least 19 recordsLinked to original sources

PubMed1999

Rescue of cyclin D1 deficiency by knockin cyclin E.

D-type cyclins and cyclin E represent two very distinct classes of mammalian G1 cyclins. We have generated a mouse strain in which the coding sequences of the cyclin D1 gene (Ccnd1) have been deleted and replaced by those of human cyclin E (CCNE). In the tissues and cells of these mice, the expression pattern of human cyclin E faithfully reproduces that normally associated with mouse cyclin D1. The replacement of cyclin D1 with cyclin E rescues all phenotypic manifestations of cyclin D1 deficiency and restores normal development in cyclin D1-dependent tissues. Thus, cyclin E can functionally replace cyclin D1. Our analyses suggest that cyclin E is the major downstream target of cyclin D1.

Animals↗

PubMed1999

Timing of cyclin E gene expression depends on the regulated association of a bipartite repressor element with a novel E2F complex.

Transient induction of the cyclin E gene in late G1 gates progression into S. We show that this event is controlled via a cyclin E repressor module (CERM), a novel bipartite repressor element located near the cyclin E transcription start site. CERM consists of a variant E2F-binding site and a contiguous upstream AT-rich sequence which cooperate during G0/G1 to delay cyclin E expression until late G1. CERM binds the protein complex CERC, which disappears upon progression through G0-G1 and reappears upon entry into the following G1. CERC disappearance correlates kinetically with the liberation of the CERM module in vivo and cyclin E transcriptional induction. CERC contains E2F4/DP1 and a pocket protein, and sediments faster than classical E2F complexes in a glycerol gradient, suggesting the presence of additional components in a novel high molecular weight complex. Affinity purified CERC binds to CERM but not to canonical E2F sites, thus displaying behavior different from known E2F complexes. In cells nullizygous for members of the Rb family, CERC is still detectable and CERM-dependent repression is functional. Thus p130, p107 and pRb function interchangeably in CERC. Notably, the CERC-CERM complex dissociates prematurely in pRb-/- cells in correspondence with the premature expression of cyclin E. Thus, we identify a new regulatory module that controls repression of G1-specific genes in G0/G1.

3T3 Cells↗

PubMed1999

Genomic distribution and gonadal mRNA expression of two human luteinizing hormone receptor exon 1 sequences in random populations.

Exon 1 of the human luteinizing hormone receptor (LHR) gene coding region exhibits at least two forms of sequence heterogeneity between 37 and 60 bp, spanning the junction of the signal peptide and the amino terminus of the mature protein. The LHR 1 differs from the LHR 2 by the insertion of 6 bp in exon 1 but is of identical sequence in the 5' flanking region. RFLP analysis of the two haplotypes within a random population of 63 individuals revealed allele frequencies of 0. 37 and 0.63 for LHR 1 and LHR 2, respectively. 94% of the samples contained at least one LHR 2 allele, whereas only 68% contained the LHR 1 allele. No gender differences were observed, and both homozygotes and heterozygotes displayed apparently normal reproduction. Reverse-transcriptase polymerase chain-reaction analyses of LHR mRNA from testes and ovaries revealed that both haplotypes are transcribed in normal individuals, with no difference in tissue specific distribution. Thus, at least two functional polymorphic forms of exon 1 coding region of the same LHR gene are present in a random human population.

Adult↗

PubMed1998

Cyclic GMP and cGMP-binding phosphodiesterase are required for interleukin-1-induced nitric oxide synthesis in human articular chondrocytes.

This study addressed the role of guanylyl cyclase (GC) and phosphodiesterase (PDE) in interleukin (IL)-1 activation of human articular chondrocytes. The GC inhibitors LY83583 and methylene blue dose-dependently inhibited IL-1-induced nitric oxide (NO) production, inducible NO synthase (iNOS) protein, and mRNA expression. These effects of GC inhibition were consistent with the rapid induction of cGMP by IL-1, which reached maximal levels after 5 min. The effects of GC inhibitors were selective as they did not reduce IL-1-induced cyclooxygenase II protein and mRNA. An inhibitor specific for soluble GC did not affect IL-1-induced NO production, and activators of soluble GC did not induce NO. However, the expression of iNOS mRNA was induced by atrial natriuretic peptide (ANP) and C-type natriuretic peptide (CNP), activators of particulate GC, indicating that particulate rather than soluble guanylyl cyclases were involved in iNOS induction. The expression of iNOS mRNA and the production of NO were induced by a slowly hydrolyzable analog of cGMP, 8-bromo-cGMP, but not by nonhydrolyzable analog, dibutyryl cGMP, suggesting that PDE rather than cGMP-dependent protein kinase mediates the cGMP effects. Chondrocytes contained extensive cGMP PDE activity. This had PDE5 biochemical features and an inhibitor profile consistent with PDE5. Furthermore, the nonisoformspecific PDE inhibitor IBMX and PDE5-specific inhibitors suppressed IL-1-induced NO release and iNOS mRNA expression. PDE5 mRNA was constitutively expressed in chondrocytes. In addition to increasing PDE5 activities, IL-1 treatment reduced the sensitivity of PDE5 to several pharmacological inhibitors by up to 50-fold. In summary, inhibitors of either GC or PDE5 prevented IL-1 induction of iNOS; IL-1 increased the rates of both cGMP generation and hydrolysis; and exogenous PDE hydrolyzable cGMP analog induced iNOS and NO. These results suggest that increased cGMP metabolic flux is sufficient to induce iNOS, and GC and PDE5 activities are required for IL-1 induction of iNOS expression via increases in coupled cGMP synthesis and hydrolysis.

1-Methyl-3-isobutylxanthine↗

PubMed1998

Nicotine-induced modulation of T Cell function. Implications for inflammation and infection.

Tobacco smoking may predispose humans to respiratory disease, and may be a compounding risk factor in HIV infection and progression to AIDS. We have demonstrated that chronic exposure of mice and rats to cigarette smoke or nicotine inhibits T cell responsiveness, which may account for the decreased antibody response to T-dependent antigens seen in these animals. This inhibition may result from aberrant antigen-mediated signaling and depletion of IP3-sensitive Ca2+ stores in nicotine-treated animals. Moreover, nicotine appears to moderate the inflammation associated with turpentine-induced sterile abscess and influenza infection. These anti-inflammatory properties of nicotine may account for longer survival of nicotine-treated than control mice lethally infected with influenza virus. However, because inflammation is required for clearance of many pathogens, nicotine-treated mice exhibit significantly higher titers of influenza virus following infection. These results offer an explanation for the higher susceptibility to some infectious diseases, but greater resistance to some inflammatory diseases among human smokers.

Abscess↗

PubMed1998

Effect of nicotine on the immune system: possible regulation of immune responses by central and peripheral mechanisms.

Nicotine (NT) treatment impairs T-cell receptor (TCR)-mediated signaling, leading to the arrest of T cells in the G1 phase of the cell cycle and inhibition of the antibody plaque-forming cell (AFC) response to sheep red blood cells (SRBC). This paper summarizes some of the previous findings related to cigarette smoke/NT and the immune response, and presents preliminary evidence suggesting that mice chronically treated with NT (0.5 mg/day/kg body weight) have a depressed inflammatory response in the turpentine-induced abscess model of inflammation. This ability of nicotine to attenuate an inflammatory response may also be the cause of reduced mortality of chronically nicotine-treated mice from acute influenza A pneumonitis. Moreover, in LEW rats, decreased anti-SRBC AFC responses were also observed after intracerebroventricular (i.c.v.) administration of relatively small concentrations of NT (28 micrograms/day/kg body weight) which, when given peripherally, did not affect the AFC response. In vitro the addition of NT to T cells increased protein tyrosine kinase (PTK) activity and intracellular Ca2+ concentration [Ca2+]i. These results support the hypothesis that NT alters immune responses by directly interacting with T cells, as well as indirectly through brain-immune interactions.

Abscess↗

PubMed1998

Neutral endopeptidase 24.11 loss in metastatic human prostate cancer contributes to androgen-independent progression.

Neutral endopeptidase 24.11 (NEP) is a cell-surface enzyme expressed by prostatic epithelial cells that cleaves and inactivates neuropeptides implicated in the growth of androgen-independent prostate cancer (PC). We report that NEP expression and catalytic activity are lost in vitro in androgen-independent but not androgen-dependent PC cell lines. In vivo, NEP protein expression is commonly decreased in cancer cells of metastatic PC specimens from patients with androgen-independent but not androgen-dependent PC. Overexpression of NEP in androgen-independent PC cells or incubation with recombinant NEP inhibits PC cell growth. Furthermore, in androgen-dependent PC cells, expression of NEP is transcriptionally regulated by androgen and decreases with androgen withdrawal. These data suggest that decreased NEP expression, common in androgen-independent PCs, is facilitated by the elimination of androgens, and that NEP loss plays an important role in the development of androgen-independent PC by allowing PC cells to use mitogenic neuropeptides as an alternate source to androgen in order to stimulate cell proliferation.

Biomarkers, Tumor↗

PubMed1998

A new set of chemotaxis homologues is essential for Myxococcus xanthus social motility.

Myxococcus xanthus cells aggregate and develop into multicellular fruiting bodies in response to starvation. A new M. xanthus locus, designated diffor defective in fruiting, was identified by the characterization of a mutant defective in fruiting body formation. Molecular cloning, DNA sequencing and sequence analysis indicate that the dif locus encodes a new set of chemotaxis homologues of the bacterial chemotaxis proteins MCPs (methyl-accepting chemotaxis proteins), CheW, CheY and CheA. The dif genes are distinct genetically and functionally from the previously identified M. xanthus frz chemotaxis genes, suggesting that multiple chemotaxis-like systems are required for the developmental process of M. xanthus fruiting body formation. Genetic analysis and phenotypical characterization indicate that the M. xanthus dif locus is required for social (S) motility. This is the first report of a M. xanthus chemotaxis-like signal transduction pathway that could regulate or co-ordinate the movement of M. xanthus cells to bring about S motility.

Amino Acid Sequence↗

PubMed1998

A DnaK homolog in Myxococcus xanthus is involved in social motility and fruiting body formation.

Myxococcus xanthus is a gram-negative soil bacterium which exhibits a complex life cycle and social behavior. In this study, two developmental mutants of M. xanthus were isolated through Tn5 transposon mutagenesis. The mutants were found to be defective in cellular aggregation as well as in sporulation. Further phenotypic characterization indicated that the mutants were defective in social motility but normal in directed cell movements. Both mutations were cloned by a transposon-tagging method. Sequence analysis indicated that both insertions occurred in the same gene, which encodes a homolog of DnaK. Unlike the dnaK genes in other bacteria, this M. xanthus homolog appears not to be regulated by temperature or heat shock and is constitutively expressed during vegetative growth and under starvation. The defects of the mutants indicate that this DnaK homolog is important for the social motility and development of M. xanthus.

Bacterial Proteins↗

PubMed1998

Chemotaxis in Borrelia burgdorferi.

Borrelia burgdorferi is a motile spirochete which has been identified as the causative microorganism in Lyme disease. The physiological functions which govern the motility of this organism have not been elucidated. In this study, we found that motility of B. burgdorferi required an environment similar to interstitial fluid (e.g., pH 7.6 and 0.15 M NaCl). Several methods were used to detect and measure chemotaxis of B. burgdorferi. A number of chemical compounds and mixtures were surveyed for the ability to induce positive and negative chemotaxis of B. burgdorferi. Rabbit serum was found to be an attractant for B. burgdorferi, while ethanol and butanol were found to be repellents. Unlike some free-living spirochetes (e.g., Spirochaeta aurantia), B. burgdorferi did not exhibit any observable chemotaxis to common sugars or amino acids. A method was developed to produce spirochete cells with a self-entangled end. These cells enabled us to study the rotation of a single flagellar bundle in response to chemoattractants or repellents. The study shows that the frequency and duration for pausing of flagella are important for chemotaxis of B. burgdorferi.

Adaptation, Physiological↗

PubMed1998

Methylation of FrzCD defines a discrete step in the developmental program of Myxococcus xanthus.

Myxococcus xanthus is a gram-negative soil bacterium which undergoes fruiting body formation during starvation. The frz signal transduction system has been found to play an important role in this process. FrzCD, a methyl-accepting taxis protein homologue, shows modulated methylation during cellular aggregation, which is thought to be part of an adaptation response to an aggregation signal. In this study, we assayed FrzCD methylation in many known and newly isolated mutants defective in fruiting body formation to determine a possible relationship between the methylation response and fruiting morphology. The results of our analysis indicated that the developmental mutants could be divided into two groups based on their ability to show normal FrzCD methylation during development. Many mutants blocked early in development, i.e., nonaggregating or abnormally aggregating mutants, showed poor FrzCD methylation. The well-characterized asg, bsg, csg, and esg mutants were found to be of this type. The defects in FrzCD methylation of these signaling mutants could be partially rescued by extracellular complementation with wild-type cells or addition of chemicals which restore their fruiting body formation. Mutants blocked in late development, i.e., translucent mounds, showed normal FrzCD methylation. Surprisingly, some mutants blocked in early development also exhibited a normal level of FrzCD methylation. The characterized mutants in this group were found to be defective in social motility. This indicates that FrzCD methylation defines a discrete step in the development of M. xanthus and that social motility mutants are not blocked in these early developmental steps.

Bacterial Proteins↗

PubMed1998

A novel human luteinizing hormone receptor gene.

A novel human luteinizing hormone receptor (LHR) gene was isolated from a human placental genomic library. This gene (Gene II) differs from that previously isolated from a lymphocyte library (Gene I) by several base changes in the 5' flanking region and the deletion of 6 nucleotides in the coding region (+55 to +60). The sequence of the exon 1 coding region of gene II conforms to the sequence of the human ovarian LHR cDNA. Furthermore, primer extension and reporter gene analyses indicate that both the Gene II promoter and the human ovarian LHR mRNA transcriptional start sites are located within the -176 bp TATA-less 5' flanking domain. Additional upstream transcriptional start sites (> -176 bp) were identified in human testicular mRNA and the human choriocarcinoma JAR cell. Restriction enzyme analysis verifies that both LHR genes are present in the human genome, and gene dosing reveals four copies of the human LHR in contrast to a single copy in the rat genome. Chromosomal mapping localizes all copies of the human LHR to the chromosome 2p16-21 loci. These studies suggest that tissue-specific LHR promoter utilization and LHR gene expression may be correlated with gene diversity.

Amino Acid Sequence↗

PubMed1996

Optimization of Long-Term Stability of Magnetic Fluids from Magnetite and Synthetic Polyelectrolytes

Nanometer-sized suspensions of magnetite (Fe3O4) stabilized with polymeric surfactants, principally poly(methacrylic acid) (PMAA), were prepared by precipitation or sonication and studied by a variety of techniques. The long-term stability of the ferrofluids made with PMAA was optimal at pH 7 (close to the measured isoelectric point of the magnetite, 5.9) and improved at all pH values with increasing surfactant concentration. With extended times of sonication the molecular weight and polydispersity of the PMAA decreased, although magnetite in the ferrofluid was not oxidized (X-ray). Particle diameters of 9-38 nm were measured by saturation magnetization, electron microscopy, and dynamic light scattering. Centrifugation at 8,000 rpm removed the magnetite from the suspensions together with roughly one molecule of PMAA for each magnetite particle in the fluid, with the average number increasing with higher concentrations and average molecular weights of the polymer.

Journal Article↗

PubMed1996

Cyclin D2 is an FSH-responsive gene involved in gonadal cell proliferation and oncogenesis.

THE D-type cyclins (D1, D2 and D3) are critical governors of the cell-cycle clock apparatus during the G1 phase of the mammalian cell cycle. These three D-type cyclins are expressed in overlapping, apparently redundant fashion in the proliferating tissues. To investigate why mammalian cells need three distinct D-type cyclins, we have generated mice bearing a disrupted cyclin D2 gene by using gene targeting in embryonic stem cells. Cyclin D2-deficient females are sterile owing to the inability of ovarian granulosa cells to proliferate normally in response to follicle-stimulating hormone (FSH), whereas mutant males display hypoplastic testes. In ovarian granulosa cells, cyclin D2 is specifically induced by FSH via a cyclic-AMP-dependent pathway, indicating that expression of the various D-type cyclins is under control of distinct intracellular signalling pathways. The hypoplasia seen in cyclin D2(-/-) ovaries and testes prompted us to examine human cancers deriving from corresponding tissues. We find that some human ovarian and testicular tumours contain high levels of cyclin D2 messenger RNA.

Animals↗

PubMed1996

Selective activation of the mitogen-activated protein kinase subgroups c-Jun NH2 terminal kinase and p38 by IL-1 and TNF in human articular chondrocytes.

Previous studies suggested that tyrosine kinase activation is an important signal transduction event in the IL-1 response of chondrocytes. The present study identifies the mitogen-activated protein (MAP) kinases extracellular signal-regulated kinase (ERK)-1 and ERK-2 as major tyrosine phosphorylated proteins in IL-1 stimulated chondrocytes. Kinase assays on immunoprecipitates with myelin basic protein as substrate showed that ERK-1 and ERK-2 activation was detectable within 5 min after IL-1 stimulation and decreased to baseline within 60 min. Analysis of other members of the MAP kinase family showed that chondrocytes also express c-Jun NH2 terminal kinase (JNK)-1, JNK-2, and p38 proteins. These kinases were time-dependently activated by IL-1. Among other chondrocyte activators tested, only TNF activated all three of the MAP kinase subgroups. JNK and p38 were not activated by any of the other cytokines and growth factors tested. However, ERK was also activated by PDGF, IGF-1, and IL-6. Phorbol 12-myristate 13-acetate, calcium ionophore, and cAMP analogues only increased ERK activity but had no significant effects on JNK or p38. These results suggest differential activation of MAP kinase subgroups by extracellular stimuli. ERK is activated in response to qualitatively diverse extracellular stimuli and various second messenger agonists. In contrast, JNK and p38 are only activated by IL-1 or TNF, suggesting that these kinases participate in the induction of the catabolic program in cartilage.

Calcium↗

PubMed1996

The genomic structure of the rat corticotropin releasing factor receptor. A member of the class II G protein-coupled receptors.

Isolation and structural characterization of the rat corticotropin releasing factor receptor (CRFR) gene was performed to determine the exon/intron organization of the coding region and the potential for splice variants. The CRFR gene contains 13 exons and 12 introns, and the positions of the exon/intron junctions are similar to those of other Class II G protein-coupled receptor genes including the parathyroid hormone and glucagon receptors. The promoter resides within 593 base pairs of the initiation codon and the major transcriptional start site at nucleotide -238. This domain does not possess a TATA box but contains multiple Sp1 and AP-2 sites upstream and downstream of the major transcriptional start site. Intron junctions were identified in the extracellular, transmembrane (TM), and cytoplasmic (C) domains of the CRFR, giving the potential for differential signal transduction by splice variants. CRFR cDNAs derived from rat Leydig cell mRNA included the pituitary Form A, which spans exons 1-13, and two splice variants with deletion of exon 3 or exons 7, 11, and 12. An evolutionary link between the intronless TM/C module of the glycoprotein hormone receptors and the intron-containing TM/C module of the CRFR is suggested by the common position of the luteinizing hormone receptor Form D alternate acceptor splice site and the CRFR intron 12.

Alternative Splicing↗

PubMed1996

Treatment of a human breast cancer xenograft with an adenovirus vector containing an interferon gene results in rapid regression due to viral oncolysis and gene therapy.

Treatment of a human breast cancer cell line (MDA-MB-435) in nude mice with a recombinant adenovirus containing the human interferon (IFN) consensus gene, IFN-con1 (ad5/IFN), resulted in tumor regression in 100% of the animals. Tumor regression occurred when virus was injected either within 24 hr of tumor cell implantation or with established tumors. However, regression of the tumor was also observed in controls in which either the wild-type virus or a recombinant virus containing the luciferase gene was used, although tumor growth was not completely suppressed. Tumor regression was accompanied by a decrease in p53 expression. Two other tumors, the human myelogenous leukemic cell line K562 and the hamster melanoma tumor RPMI 1846, also responded to treatment but only with ad5/IFN. In the case of K562 tumors, there was complete regression of the tumor, and tumors derived from RPMI 1846 showed partial regression. We propose that the complete regression of the breast cancer with the recombinant virus ad5/IFN was the result of two events: viral oncolysis in which tumor cells are being selectively lysed by the replication-competent virus and the enhanced effect of expression of the IFN-con1 gene. K562 and RPMI 1846 tumors regressed only as a result of IFN gene therapy. This was confirmed by in vitro analysis. Our results indicate that a combination of viral oncolysis with a virus of low pathogenicity, itself resistant to the effects of IFN and IFN gene therapy, might be a fruitful approach to the treatment of a variety of different tumors, in particular breast cancers.

Adenoviruses, Human↗

PubMed1996

Effects of nicotine on the immune response. II. Chronic nicotine treatment induces T cell anergy.

Previously, we have shown that both T and B lymphocytes from chronically nicotine-treated (NT) animals exhibit tolerance to activation by Ags (ligation of Ag receptors), as indicated by their decreased ability to mobilize intracellular calcium and, at least in T cells, arrest of cells in the G0/G1 phase of the cell cycle. Herein, we demonstrate that NT T cells significantly lose their ability to up-regulate inositol trisphosphate synthesis in response to TCR ligation or nonspecific activation of G proteins by AIF-4. However, increases in cAMP concentrations of T cells following activation of G protein-sensitive adenylate cyclase by cholera or pertussis toxin were not significantly affected by the nicotine treatment. Interestingly, compared with control T cells, the background levels of inositol trisphosphate were significantly elevated in NT T cells, indicating some degree of activation in these cells. This inference was further supported by observations that naive T cells from NT animals exhibit tyrosine phosphorylation of several substrates, including phospholipase C-gamma1, which were either absent or underphosphorylated in unstimulated control T cells. Moreover, when, after 4-wk nicotine treatment, nicotine pumps were removed and serum cotinine levels fell to background, inhibition of the Ab-forming cells and Ca2+ responses continued for at least 2 more wk. These results suggest that chronic in vivo nicotine exposure leads to T cell anergy and may contribute to nicotine/cigarette smoke-induced immunosuppression.

Aluminum Compounds↗