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Biomedical subjects

Q Chen

Publications and source records attributed to Q Chen.

At least 379 records · Page 21Linked to original sources
PubMed1998

The different functions of BglF, the E. coli beta-glucoside permease and sensor of the bgl system, have different structural requirements.

The Escherichia coli BglF protein (EIIbgl) is an Enzyme II (EII) of the phosphoenolpyruvate-dependent phosphotransferase system (PTS) which catalyses transport and phosphorylation of beta-glucosides. In addition to its transport function, BglF serves as a beta-glucoside sensor which reversibly phosphorylates BglG, the transcription regulator of the bgl operon. Like many other PTS sugar permeases, the BglF protein is composed of three discrete functional and structural domains: IIAbgl and IIBbgl, which are hydrophilic, and IICbgl, which is hydrophobic. The domains of BglF are covalently linked to one another in the order BCA. The IIAbgl domain contains the first phosphorylation site, which accepts a phosphoryl group from the general PTS protein HPr and delivers it to the second phosphorylation site, located in the IIBbgl domain. This second site can deliver the phosphoryl group either to a beta-glucoside or to BglG. To elucidate the mechanism by which such different substrates can be phosphorylated by the same active site, we decided to try to separate the different phosphorylation activities catalyzed by BglF. To this end we rearranged the BglF domains and constructed IICBAbgl (scrambled-BglF). Scrambled-BglF behaved like wild-type BglF in its ability to be phosphorylated and to phosphorylate BglG in vitro and in vivo. However, it could not catalyze phosphorylation of beta-glucosides in vitro nor their phosphotransfer in vivo, and it could not catalyze BglG dephosphorylation in vitro or in vivo. Therefore, the two reactions induced by beta-glucosides, sugar phosphorylation and BglG dephosphorylation, seem to require a specific domain organization: IIBbgl should precede IICbgl. The order of the B and C domains is irrelevant for BglG phosphorylation, which occurs in the absence of beta-glucosides. Because the domain order affects the way that the domains are able to interact, our results suggest that catalysis of the sugar-induced functions depends on specific interactions between IIBbgl and IICbgl. In light of the previous assumption that domain order in EIIs is immaterial for their function, the finding that the order of the domains is important for the function of BglF as a sugar phosphotransferase raises two possibilities: (a) BglF differs from other EIIs in this regard; (b) BglF represents a subgroup of EIIs in which the requirement for a specific domain order correlates with the ability to transport a set of structurally related sugars.

Amino Acid Sequence↗
PubMed1998

[The food habit and its affecting factors of preschool children in Guangzhou].

A survey of nutritional knowledge-attitude-practice was conducted among 1300 preschool children, their parents, 203 teachers and nurses in Kindergarten in 1997. Results showed that the food habit of preschool children was unsuitable. Most preschool children rejected eating animal liver, peanut product and animal blood, but often ate snacks. From statistic analysis among affecting factors, the mother's and teachers' nutritional knowledge and the food habit of parents had significant effect on food habit of preschool children. The family income did not significantly affect food habit of preschool children. Moreover, there was little nutrition education involved in the preschool children courses in 4 kindergartens. The result suggested that the nutrition education plan should be done.

Adult↗
PubMed1998

Developmental selection of var gene expression in Plasmodium falciparum.

The protozoan Plasmodium falciparum causes lethal malaria. Adhesion of erythrocytes infected with P. falciparum to vascular endothelium and to uninfected red blood cells (rosetting) may be involved in the pathogenesis of severe malaria. The binding is mediated by the antigenically variant erythrocyte-membrane-protein-1 (PfEMP-1), which is encoded by members of the P. falciparum var gene family. The control of expression and switching of var genes seems to lack resemblance to mechanisms operating in variant gene families of other microbial pathogens. Here we show that multiple, distinct var gene transcripts (about 24 or more) can be detected by reverse transcription and polymerase chain reaction in bulk cultures of the rosetting parasite FCR3S1.2, despite the adhesive homogeneity of the cultures. We also detected several var transcripts in single erythrocytes infected with a ring-stage parasite of FCR3S1.2, and found that different var genes are transcribed simultaneously from several chromosomes in the same cell. In contrast, we detected only one var transcript, FCR3S1.2 var-1, which encodes the rosetting PfEMP-1 protein, in individual rosette-adhesive trophozoite-infected cells, and we found only one PfEMP-1 type at the erythrocyte surface by labelling with 125iodine and immunoprecipitation. We conclude that a single P. falciparum parasite simultaneously transcribes multiple var genes but, through a developmentally regulated process, selects only one PfEMP-1 to reach the surface of the host cell.

Adult↗
PubMed1998

BglF, the sensor of the bgl system and the beta-glucosides permease of Escherichia coli: evidence for dimerization and intersubunit phosphotransfer.

The Escherichia coli BglF protein, also designated EIIbgl, is an enzyme II of the phosphoenolpyruvate-dependent phosphotransferase system (PTS) that catalyzes transport and phosphorylation of beta-glucosides. In addition, BglF has the ability, unusual for an EII, to regulate the activity of a transcriptional regulator, BglG, by phosphorylating and dephosphorylating it according to beta-glucoside availability. Together, BglF and BglG constitute a novel sensory system. The membrane-bound sensor, BglF, has two phosphorylation sites: site 1 accepts a phosphoryl group from HPr and delivers it to site 2; site 2 delivers the phosphoryl group either to beta-glucosides or to BglG. Here, we provide several lines of evidence for the dimerization of BglF and for the occurrence of productive intersubunit phosphotransfer within the BglF dimers. (1) Two inactive BglF mutant proteins, one lacking phosphorylation site 1 and the other lacking site 2, complement one another to allow beta-glucoside utilization by bglF strains. (2) The pairs of mutant proteins complement one another in regulating BglG activity as a transcriptional antiterminator in vivo. (3) Only when they are present in the same membrane preparation do the mutant protein pairs efficiently transfer the phosphoryl group from HPr to beta-glucosides and to BglG in vitro. (4) Gentle extraction of cellular proteins followed by SDS-PAGE reveals the existence of BglF homodimers. A portion of the phosphorylated form of BglF can also be extracted from the membrane as a dimer. Dimerization is mediated by the membrane-bound IICbgl domain, as indicated by the dimerization of IICbgl by itself and of BglF derivatives that contain this domain. Since dimers persist in the presence of a reducing agent, they are apparently not held together by disulfide bonds. Rather, BglF dimerization might involve hydrophobic interactions between residues in the membrane-spanning domain. In addition, we show that BglF dimerization is not modulated by beta-glucosides and is therefore not part of the mechanism that diverts the phosphoryl group away from BglG to the transported sugar upon addition of beta-glucosides to the growth medium.

Biosensing Techniques↗
PubMed1998

[A molecular variant of angiotensinogen gene is associated with myocardial infarction in Chinese].

OBJECTIVE: A molecular variant of the angiotensinogen(AGT) gene with threonine instead of methionine at position 235(i.e.,with M235T polymor- phism) has been shown to be associated with essential hypertension, preeclampsia, coronary atherosclerosis, coronary heart disease(CHD), myocardial infarction(MI) in Caucasians and in Japanese. The purpose of the present study was to assess whether the M235T polymorphism was associated with MI in a Chinese population. METHODS: M235T polymorphism in exon 2 of AGT gene was determined by polymerase chain reaction(PCR) and restriction endonuclease analysis in a study of 57 patients with MI and 76 non-CHD individuals as control. RESULTS: The frequencies of T235 allele(0.82) and 235TT genotype (0.70) in the MI group were higher than those in the control subjects(0.63 and 0.42 respectively, P=0.013 and P<0.025). AGT gene 235TT genotype was at significantly increased risk of MI(odds ratio 3.65, P=0.016) in analysis adjusted for several main CHD risk factors. CONCLUSION: There is a significant association between AGT gene 235TT genotype and MI, this genotype might be an independent risk for MI in Chinese population.

Aged↗
PubMed1998

Increased transcription decreases the spontaneous mutation rate at the thymidine kinase locus in human cells.

Transcription increases DNA repair efficiency and modulates the distribution of certain types of DNA damage. Furthermore, increased transcription level stimulates spontaneous mutation rate in yeast. We explored whether transcription level affects spontaneous mutation rate in human cells. We first developed two thymidine kinase (tk) inducible human cell lines using the Gal4-Estrogen receptor system. In our TK6i-G3 and G9 tk heterozygous cell lines, the active tk allele is linked to an inducible promoter element. Tk mRNA is induced following treatment with estrogen. Spontaneous mutation rate was significantly decreased in human cell lines after induction in contrast to the report in yeast. Thus, humans may have evolved different or additional mechanisms to deal with transcription related spontaneous mutagenesis.

B-Lymphocytes↗
PubMed1998

NF-kappa B is activated and ICAM-1 gene expression is upregulated during reoxygenation of human brain endothelial cells.

Reperfusion injury is mediated, in part, by the upregulated expression of genes in microvascular endothelial cells that encode for inflammatory cytokines and adhesion molecules. The redox-regulated transcription factor, nuclear factor kappa B (NF-kappaB), may play a major role in the induced expression of these genes. In this study we use cultured human brain microvascular endothelial cells (HBMEC) to investigate whether reoxygenation of hypoxic HBMEC results in the activation of NF-kappaB and the upregulation of the adhesion molecule, ICAM-1. When HBMEC were subjected to hypoxia followed by reoxygenation but not hypoxia alone, an NF-kappaB complex composed of p65 and p50 Rel proteins was rapidly activated within 15-30 min. Four hours later, expression of the ICAM-1 gene was significantly upregulated. The antioxidant pyrrolidine dithiocarbamate and the proteasome inhibitor, n-Tosyl-Phe-chloromethyl ketone, blocked both the activation of NF-kappaB and the upregulation of the ICAM-1 gene. These results indicate that NF-kappaB is activated in HBMEC by reoxygenation and may play a significant role in the upregulation of the ICAM-1 gene. Agents which inhibit NF-kappaB activation may be potential therapeutic agents in acute ischemic stroke.

Brain↗
PubMed1998

The role of type I and type II tumor necrosis factor (TNF) receptors in the ability of TNF-alpha to transduce a proliferative signal in the human megakaryoblastic leukemic cell line Mo7e.

We investigated the effects of tumor necrosis factor (TNF) alpha on the human megakaryocytic leukemic cell lines Mo7e, Meg-01, and Dami/HEL. Our data show that both type I and type II TNF receptors (TNF-RI and TNF-RII) are expressed on all of these cells, and TNF-alpha significantly stimulates the proliferation of growth factor-dependent Mo7e cells but not of Meg-01 or Dami/HEL cells, which grow in a factor-independent manner. TNF-alpha serves predominantly as a mitogen for Mo7e cell proliferation and does not induce Mo7e cell differentiation. Coincubation with both TNF-alpha and anti-TNF-alpha neutralizing antibody completely abolishes the TNF-alpha-induced proliferation of Mo7e cells. In bioassays, there is no detectable level of other stimulatory cytokines in conditioned medium from Mo7e cells previously stimulated by TNF-alpha, implying that the stimulatory effect of TNF-alpha on Mo7e cells is derived from the direct action of TNF-alpha rather than via the induction of secondary cytokines by TNF-alpha. Flow cytometric studies demonstrated that TNF-alpha binds to Mo7e cells that have been pretreated with either anti-TNF-RI or anti-TNF-RII neutralizing antibody, but TNF-alpha does not bind to cells pre-exposed to both receptor antibodies. However, the incubation of Mo7e cells with either TNF-RI or TNF-RII neutralizing antibodies or with either soluble TNF-RI or TNF-RII inhibits TNF-alpha-induced cell proliferation, indicating the requirement of interactions with both TNF receptors for the mitogenic activity of TNF-alpha. Furthermore, our data suggest that an alternative signaling pathway may be involved in TNF-alpha-induced Mo7e cell proliferation, because the common mitogen-activated protein kinase (MAPK) and signal transducer and activator of transcription (STAT) signaling pathways activated by other cytokines that induce Mo7e cell proliferation are not activated by TNF-alpha.

Cell Division↗
PubMed1998

New mutations in the KVLQT1 potassium channel that cause long-QT syndrome.

BACKGROUND: Long-QT syndrome (LQTS) is an inherited cardiac arrhythmia that causes sudden death in young, otherwise healthy people. Four genes for LQTS have been mapped to chromosome 11p15.5 (LQT1), 7q35-36 (LQT2), 3p21-24 (LQT3), and 4q25-27 (LQT4). Genes responsible for LQT1, LQT2, and LQT3 have been identified as cardiac potassium channel genes (KVLQT1, HERG) and the cardiac sodium channel gene (SCN5A). METHODS AND RESULTS: After studying 115 families with LQTS, we used single-strand conformation polymorphism (SSCP) and DNA sequence analysis to identify mutations in the cardiac potassium channel gene, KVLQT1. Affected members of seven LQTS families were found to have new, previously unidentified mutations, including two identical missense mutations, four identical splicing mutations, and one 3-bp deletion. An identical splicing mutation was identified in affected members of four unrelated families (one Italian, one Irish, and two American), leading to an alternatively spliced form of KVLQT1. The 3-bp deletion arose de novo and occurs at an exon-intron boundary. This results in a single base deletion in the KVLQT1 cDNA sequence and alters splicing, leading to the truncation of KVLQT1 protein. CONCLUSIONS: We have identified LQTS-causing mutations of KVLQT1 in seven families. Five KVLQT1 mutations cause the truncation of KVLQT1 protein. These data further confirm that KVLQT1 mutations cause LQTS. The location and character of these mutations expand the types of mutation, confirm a mutational hot spot, and suggest that they act through a loss-of-function mechanism or a dominant-negative mechanism.

Case-Control Studies↗
PubMed1998

The upstream regulatory regions of the hepatocyte growth factor gene promoter are essential for its expression in transgenic mice.

To understand the molecular mechanisms of hepatocyte growth factor (HGF) gene transcription in vivo, we report the generation and characterization of transgenic mice harboring various lengths of the mouse HGF promoter linked to the chloramphenicol acetyltransferase reporter gene. Analysis of different tissues of the transgenic mouse lines having the 2.7-kilobase (kb) promoter construct revealed a pattern of reporter gene expression in embryonic and adult tissues that paralleled that of endogenous HGF gene expression. A similar expression pattern was observed in the 0.7-kb transgenic lines. However, in contrast to in vitro data, no promoter activity was detected in four independent transgenic lines harboring the 0.1-kb construct. Akin to the activity of the endogenous HGF gene, which is induced in the liver, lung, and spleen in response to 70% partial hepatectomy, the reporter gene driven by the 2.7-kb promoter construct was strongly induced, whereas that driven by the 0.7-kb promoter construct was modestly induced in these organs after partial hepatectomy. Together, these data suggest that the region between -0.1 and -0.7 kb of the HGF gene promoter is essential to drive its expression in vivo and that additional upstream sequences located between -0.7 and -2.7 kb are also necessary for its maximum inducibility in response to cues that stimulate tissue growth and regeneration.

Animals↗
PubMed1998

Genetic basis and molecular mechanism for idiopathic ventricular fibrillation.

Ventricular fibrillation causes more than 300,000 sudden deaths each year in the USA alone. In approximately 5-12% of these cases, there are no demonstrable cardiac or non-cardiac causes to account for the episode, which is therefore classified as idiopathic ventricular fibrillation (IVF). A distinct group of IVF patients has been found to present with a characteristic electrocardiographic pattern. Because of the small size of most pedigrees and the high incidence of sudden death, however, molecular genetic studies of IVF have not yet been done. Because IVF causes cardiac rhythm disturbance, we investigated whether malfunction of ion channels could cause the disorder by studying mutations in the cardiac sodium channel gene SCN5A. We have now identified a missense mutation, a splice-donor mutation, and a frameshift mutation in the coding region of SCN5A in three IVF families. We show that sodium channels with the missense mutation recover from inactivation more rapidly than normal and that the frameshift mutation causes the sodium channel to be non-functional. Our results indicate that mutations in cardiac ion-channel genes contribute to the risk of developing IVF.

Action Potentials↗
PubMed1998

Fenamates protect neurons against ischemic and excitotoxic injury in chick embryo retina.

Three fenamates (flufenamate, meclofenamate and mefenamate) were examined for their protective effect on neurons under ischemic (glucose/oxygen deprivation) or excitotoxic conditions, using the isolated retina of chick embryo as a model. Retinal damage was evaluated by histology and lactate dehydrogenase assay. Whole-cell recording was used to examine the direct effect of the fenamates on glutamate receptor-mediated currents. The fenamates protected the retina against the ischemic or excitotoxic insult. Part of the neuroprotection by the fenamates derived from inhibition of N-methyl-D-aspartate receptor-mediated currents. However, kainate receptor-mediated currents were not blocked by the fenamates, which nonetheless reduced kainate receptor-mediated retinal damage. Our results raise the possibility that fenamates may serve as lead structures in the development of novel therapeutic agents against brain ischemia.

Animals↗
PubMed1998

Sequence of a 131-kb region of 5q13.1 containing the spinal muscular atrophy candidate genes SMN and NAIP.

The spinal muscular atrophies (SMA), which are characterized by motor neuron loss and progressive paralysis, are among the most common autosomal recessive disorders. The SMA region of chromosome 5q13.1 is distinguished by variable amplification of genomic sequence incorporating a number of genes and pseudogenes. Recently, two SMA candidate genes mapping to this area were identified: survival motor neuron (SMN) and neuronal apoptosis inhibitory protein (NAIP). The telomeric copy of SMN (SMNtel) is deleted in over 95% of cases of SMA, with NAIP deletions primarily seen in type I SMA. We present here 131 kb of genomic sequence from 5q13.1 incorporating both NAIP and SMNtel in addition to revisions of the original NAIP cDNA sequence. The Alu-rich NAIP-SMNtel interval contains the microsatellite polymorphisms that are deleted in as many as 80% of type I SMA chromosomes, focusing attention on this region in the pathogenesis of type I SMA.

Amino Acid Sequence↗
PubMed1998

Analysis of the acyl-CoAs that accumulate during the peroxisomal beta-oxidation of arachidonic acid and 6,9,12-octadecatrienoic acid.

The biosynthesis of 4,7,10,13,16-22:5 and 4,7,10,13,16,19-22:6 requires that when 6,9,12,15,18-24:5 and 6,9,12,15,18,21-24:6 are produced in microsomes they must move to peroxisomes for partial beta-oxidation. When the 24-carbon acids were incubated with peroxisomes, 22-carbon acids with their first double bond at position 4 accumulated as did those with their first two double bonds at the 2-trans-4-cis-positions (D. L. Luthria, S. B. Mohammed, and H. Sprecher, J. Biol. Chem. 271, 16020-16025, 1996; and B. S. Mohammed, D. L. Luthria, S. P. Baykousheva, and H. Sprecher, Biochem. J., 326, 425-430, 1997). In the study reported here we analyzed the acyl-CoAs that accumulated when peroxisomes were incubated with 5,8,11,14-20:4 and 6,9,12-18:3, a metabolite that would be produced via one cycle of arachidonate degradation via the pathway requiring both NADPH-dependent 2,4-dienoyl-CoA reductase and delta 3,5, delta 2,4-dienoyl-CoA isomerase. With both substrates the acyl-CoAs of 2-trans-4-10:2, 4-10:1, 2-trans-4,7,10-16:4, and 4,7,10-16:3 accumulated. These results further establish that the reductase catalyzes a control step in the peroxisomal degradation of unsaturated fatty acids. It was not possible to detect any 18- or 12-carbon acyl-CoA when arachidonate was the substrate, nor did any 12-carbon catabolite accumulate from 6,9,12-18:3. The fractional amount of 5,8-14:2 and arachidonate catabolized via the pathway using only the enzymes of saturated fatty acid degradation versus the pathway that also uses the reductase and the isomerase could thus not be estimated.

Acyl Coenzyme A↗
PubMed1998

Identification of Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) as the rosetting ligand of the malaria parasite P. falciparum.

Severe Plasmodium falciparum malaria is characterized by excessive sequestration of infected and uninfected erythrocytes in the microvasculature of the affected organ. Rosetting, the adhesion of P. falciparum-infected erythrocytes to uninfected erythrocytes is a virulent parasite phenotype associated with the occurrence of severe malaria. Here we report on the identification by single-cell reverse transcriptase PCR and cDNA cloning of the adhesive ligand P. falciparum erythrocyte membrane protein 1 (PfEMP1). Rosetting PfEMP1 contains clusters of glycosaminoglycan-binding motifs. A recombinant fusion protein (Duffy binding-like 1-glutathione S transferase; Duffy binding-like-1-GST) was found to adhere directly to normal erythrocytes, disrupt naturally formed rosettes, block rosette reformation, and bind to a heparin-Sepharose matrix. The adhesive interactions could be inhibited with heparan sulfate or enzymes that remove heparan sulfate from the cell surface whereas other enzymes or similar glycosaminoglycans of a like negative charge did not affect the binding. PfEMP1 is suggested to be the rosetting ligand and heparan sulfate, or a heparan sulfate-like molecule, the receptor both for PfEMP1 binding and naturally formed erythrocyte rosettes.

Amino Acid Sequence↗
PubMed1998

Factor VII deficiency caused by a structural variant N57D of the first epidermal growth factor domain.

We have previously described a kindred with factor VII (FVII) deficiency whose members exhibited reduced procoagulant activity relative to FVII antigen concentration. In this report, the molecular genetic basis of the FVII defect has been determined to be a heterozygous substitution of Asp for Asn at position 57 in the first epidermal growth factor (EGF) domain. Recombinant FVII (N57D) cDNA was created by site-directed mutagenesis and transiently expressed in human 293 cells. The transfected cells synthesized an immunoprecipitable protein with an apparent molecular weight of 50 kD. Quantitation of expression by FVII enzyme-linked immunosorbent assay indicated that mutant protein yields were consistently low, typically 10% to 30% of wild-type FVII. FVII (N57D) protein did not accumulate intracellularly, and Northern blot analysis indicated equivalent FVII mRNA levels in 293 cells expressing either wild-type FVII or FVII (N57D). Secreted FVII (N57D) protein did not bind tissue factor, exhibited no procoagulant activity, and failed to bind a conformation-dependent monoclonal antibody specific for the first EGF domain of FVII. Molecular modeling of the first EGF domain of FVII predicted that the N57D amino acid substitution would disrupt tertiary bonding structure. We conclude that the N57D mutation affects folding of the first EGF domain of FVII resulting in decreased cellular secretion of a mutant FVII molecule, which is unable to bind tissue factor and is therefore biologically inactive.

Animals↗
PubMed1998

EPISTAR MRI: multislice mapping of cerebral blood flow.

A method is described for multislice EPISTAR that perfectly compensates magnetization transfer effects. Inflowing arterial spins are labeled with a 360 degrees adiabatic pulse. Two control tags are applied sequentially at the same location as the labeling pulse, each with a 180 degrees adiabatic pulse so the total RF irradiation, frequency shift, and bandwidth of the labeling and control pulses are identical. Therefore, magnetization transfer effects are the same as for the labeling pulse and cancel with image subtraction for all slices. The method also eliminates tagging of venous spins and concern about asymmetric magnetization transfer effects.

Adult↗