Biomedical subjects

Q Sun

Publications and source records attributed to Q Sun.

At least 127 records · Page 7Linked to original sources

PubMed1998

Developmental changes in expression of ion currents accompany maturation of locomotor pattern in frog tadpoles.

1. The K+ currents of spinal neurons acutely dissociated from Xenopus larvae were studied and compared with those of neurons dissociated from Xenopus embryos. 2. The density of total outward current in the larval and embryonic neurons remained the same from stage 37/38 to stage 42. 3. Almost all neurons at stage 42 expressed a fast activating Ca2+-dependent K+ current (IKCa) that was largely absent from embryonic neurons. Whereas IKCa became larger and more prevalent during development, the delayed rectifier K+ currents were down-regulated. 4. About 53 % of IKCa was selectively blocked by iberiotoxin which had no effect on the delayed rectifier K+ currents or the K+ currents of embryonic neurons. 5. The firing properties of neurons isolated from embryos were unchanged by iberiotoxin. However, the toxin greatly increased the frequency of firing in larval neurons. 6. Iberiotoxin extended the duration of ventral root bursts during fictive swimming in larvae at stages 41 and 42 but had no effect at stage 40. The progressive expression of IKCa thus contributed to burst termination. 7. We have found that changes in expression of outward current closely correlate with the maturation of the motor pattern during development. At a time when the motor pattern has a need for a burst-terminating mechanism, the larval neurons express a channel with properties appropriate for such a role.

Aging↗

PubMed1998

Identification of a Cys motif in the common beta chain of the interleukin 3, granulocyte-macrophage colony-stimulating factor, and interleukin 5 receptors essential for disulfide-linked receptor heterodimerization and activation of all three receptors.

The human interleukin 3 (IL-3) and granulocyte-macrophage colony-stimulating factor (GM-CSF) receptors undergo covalent dimerization of the respective specific alpha chains with the common beta subunit (betac) in the presence of the cognate ligand. We have now performed alanine substitutions of individual Cys residues in betac to identify the Cys residues involved and their contribution to activation of the IL-3, GM-CSF, and IL-5 receptors. We found that substitution of Cys-86, Cys-91, and Cys-96 in betac but not of Cys-100 or Cys-234 abrogated disulfide-linked IL-3 receptor dimerization. However, although Cys-86 and Cys-91 betac mutants retained their ability to form non-disulfide-linked dimers with IL-3Ralpha, substitution of Cys-96 eliminated this interaction. Binding studies demonstrated that all betac mutants with the exception of C96A supported high affinity binding of IL-3 and GM-CSF. In receptor activation experiments, we found that betac mutants C86A, C91A, and C96A but not C100A or C234A abolished phosphorylation of betac in response to IL-3, GM-CSF, or IL-5. These data show that although Cys-96 is important for the structural integrity of betac, Cys-86 and Cys-91 participate in disulfide-linked receptor heterodimerization and that this linkage is essential for tyrosine phosphorylation of betac. Sequence alignment of betac with other cytokine receptor signaling subunits in light of these data shows that Cys-86 and Cys-91 represent a motif restricted to human and mouse beta chains, suggesting a unique mechanism of activation utilized by the IL-3, GM-CSF, and IL-5 receptors.

Amino Acid Sequence↗

PubMed1998

IL-3, IL-5, granulocyte-macrophage colony-stimulating factor receptor alpha-subunit, and common beta-subunit expression by peripheral leukocytes and blood dendritic cells.

BACKGROUND: IL-3, IL-5, and granulocyte-macrophage colony-stimulating factor (GM-CSF) receptors consist of cytokine-specific alpha-subunits, which associate with a shared signalling common beta-subunit (beta(c)) to form a high-affinity complex. The expression of IL-3, IL-5, and GM-CSF is upregulated in atopic inflammation, and these cytokines are thought to contribute to pathology through mechanisms that include eosinophil activation. OBJECTIVE: We sought to examine the distribution of receptor expression between cells relevant to allergic inflammation from individual subjects and to compare atopic and nonatopic individuals. METHODS: Peripheral blood was obtained from atopic and nonatopic volunteers. Cytokine-receptor expression was examined by flow cytometry with monoclonal antibodies specific for alpha-subunits and beta(c) in combination with phenotypic markers for eosinophils, basophils, neutrophils, dendritic cells, monocytes, and T cells. RESULTS: Using a ligand-independent system, we confirmed the cellular distribution of IL-5Ralpha, IL-3Ralpha, and GM-CSFRalpha. IL-3Ralpha and GM-CSFRalpha were detected on high-affinity IgE receptor blood dendritic cells. Beta(c) expression was detected on basophils, eosinophils, neutrophils, and, at low levels, on monocytes and dendritic cells. There was intense staining of basophils for IL-3Ralpha relative to IL-5Ralpha, GM-CSFRalpha, and beta(c), whereas eosinophil-staining intensity was similar for IL-3Ralpha, IL-5Ralpha, GM-CSFRalpha, and beta(c). There were no significant differences between atopic and nonatopic subjects in cytokine-receptor staining. CONCLUSION: IL-3Ralpha and GM-CSRalpha are shown on a newly defined population of Fc(epsilon)RI-high dendritic cells. The intense staining of basophils for IL-3Ralpha, relative to that of IL-5Ralpha and GM-CSFRalpha, is in contrast to eosinophils from the same subjects and may explain the higher sensitivity of basophils to IL-3 compared with IL-5 and GM-CSF. We found no evidence for downregulation of receptor expression in atopic compared with nonatopic subjects, suggesting that these receptors remain accessible as potential targets for therapeutic intervention in atopic allergic disease.

Adolescent↗

PubMed1998

Quantitative structure-activity relationships on 5-substituted terbenzimidazoles as topoisomerase I poisons and antitumor agents.

Several 5-substituted terbenzimidazoles were synthesized and evaluated as mammalian topoisomerase I poisons and for cytotoxicity against a human lymphoblastoma cell line, RPMI-8402. No correlation was observed between topoisomerase I poisoning activity and the Hansch pi value or the sigma meta and sigma para values associated with each substituent. These data suggest that electronic effects and relative lipophilicity of substituents at the 5-position of these terbenzimidazoles do not have a significant effect upon intrinsic topoisomerase I poisoning activity. There was, however, a good correlation between the relative pi values for the various substituents evaluated and cytotoxic activity. Experimentally determined log P values did not correlate well with either cytotoxicity or pi values. Capacity factors (log k') as determined by high pressure liquid chromatography did correlate well with the pi values of varied substituents and cytotoxicity. These data indicate that the relative lipophilic activity of substituents at the 5-position of these terbenzimidazoles can strongly influence relative cytotoxic activity.

Antineoplastic Agents↗

PubMed1998

Cell cycle arrest in Era GTPase mutants: a potential growth rate-regulated checkpoint in Escherichia coli.

Era is a low-molecular-weight GTPase essential for Escherichia coli viability. The gene encoding Era is found in the rnc operon, and the synthesis of both RNase III and Era increases with growth rate. Mutants that are partially defective in Era GTPase activity or that are reduced in the synthesis of wild-type Era become arrested in the cell cycle at the predivisional two-cell stage. The partially defective Era GTPase mutation (era1) suppresses several temperature-sensitive lethal alleles that affect chromosome replication and chromosome partitioning but not cell division. Our results suggest that Era plays an important role in cell cycle progression at a specific point in the cycle, after chromosome partitioning but before cytokinesis. Possible functions for Era in cell cycle progression and the initiation of cell division are discussed.

Amino Acid Sequence↗

PubMed1998

Assembly of the FtsZ ring at the central division site in the absence of the chromosome.

The FtsZ ring assembles between segregated daughter chromosomes in prokaryotic cells and is essential for cell division. To understand better how the FtsZ ring is influenced by chromosome positioning and structure in Escherichia coli, we investigated its localization in parC and mukB mutants that are defective for chromosome segregation. Cells of both mutants at non-permissive temperatures were either filamentous with unsegregated nucleoids or short and anucleate. In parC filaments, FtsZ rings tended to localize only to either side of the central unsegregated nucleoid and rarely to the cell midpoint; however, medial rings reappeared soon after switching back to the permissive temperature. Filamentous mukB cells were usually longer and lacked many potential rings. At temperatures permissive for mukB viability, medial FtsZ rings assembled despite the presence of apparently unsegregated nucleoids. However, a significant proportion of these FtsZ rings were mislocalized or structurally abnormal. The most surprising result of this study was revealed upon further examination of FtsZ ring positioning in anucleate cells generated by the parC and mukB mutants: many of these cells, despite having no chromosome, possessed FtsZ rings at their midpoints. This discovery strongly suggests that the chromosome itself is not required for the proper positioning and development of the medial division site.

Bacterial Proteins↗

PubMed1998

Localization and function of early cell division proteins in filamentous Escherichia coli cells lacking phosphatidylethanolamine.

Escherichia coli cells that contain the pss-93 null mutation are completely deficient in the major membrane phospholipid phosphatidylethanolamine (PE). Such cells are defective in cell division. To gain insight into how a phospholipid defect could block cytokinesis, we used fluorescence techniques on whole cells to investigate which step of the cell division cycle was affected. Several proteins essential for early steps in cytokinesis, such as FtsZ, ZipA, and FtsA, were able to localize as bands to potential division sites in pss-93 filaments, indicating that the generation and localization of potential division sites was not grossly affected by the absence of PE. However, there was no evidence of constriction at most of these potential division sites. FtsZ and green fluorescent protein (GFP) fusions to FtsZ and ZipA often formed spiral structures in these mutant filaments. This is the first report of spirals formed by wild-type FtsZ expressed at normal levels and by ZipA-GFP. The results suggest that the lack of PE may affect the correct interaction of FtsZ with membrane nucleation sites and alter FtsZ ring structure so as to prevent or delay its constriction.

Bacterial Proteins↗

PubMed1998

Localization of cell division protein FtsK to the Escherichia coli septum and identification of a potential N-terminal targeting domain.

Escherichia coli cell division protein FtsK is a homolog of Bacillus subtilis SpoIIIE and appears to act late in the septation process. To determine whether FtsK localizes to the septum, we fused three N-terminal segments of FtsK to green fluorescent protein (GFP) and expressed them in E. coli cells. All three segments were sufficient to target GFP to the septum, suggesting that as little as the first 15% of the protein is a septum-targeting domain. Localized fluorescence was detectable only in cells containing a visible midcell constriction, suggesting that FtsK targeting normally occurs only at a late stage of septation. The largest two FtsK-GFP fusions were able at least partially to complement the ftsK44 mutation in trans, suggesting that the N- and C-terminal domains are functionally separable. However, overproduction of FtsK-GFP resulted in a late-septation phenotype similar to that of ftsK44, with fluorescent dots localized at the blocked septa, suggesting that high levels of the N-terminal domain may still localize but also inhibit FtsK activity. Interestingly, under these conditions fluorescence was also sometimes localized as bands at potential division sites, suggesting that FtsK-GFP is capable of targeting very early. In addition, FtsK-GFP localized to potential division sites in cephalexin-induced and ftsI mutant filaments, further supporting the idea that FtsK-GFP can target early, perhaps by recognizing FtsZ directly. This hypothesis was supported by the failure of FtsK-GFP to localize in ftsZ mutant filaments. In ftsK44 mutant filaments, FtsA and FtsZ were usually localized to potential division sites between the blocked septa. When the ftsK44 mutation was incorporated into the FtsK-GFP fusions, localization to midcell ranged between very weak and undetectable, suggesting that the FtsK44 mutant protein is defective in targeting the septum.

Amino Acid Substitution↗

PubMed1998

FtsZ dynamics during the division cycle of live Escherichia coli cells.

The dynamics and assembly of bacterial cell division protein FtsZ were monitored in individual, growing and dividing Escherichia coli cells in real time by microculture of a merodiploid strain expressing green fluorescent protein (GFP)-tagged FtsZ. Cells expressing FtsZ-GFP at levels less than or equivalent to that of wild-type FtsZ were able to grow and divide over multiple generations, with their FtsZ rings visualized by fluorescence. During the late stages of cytokinesis, which constituted the last one-fourth of the cell cycle, the lumen of the FtsZ ring disappeared as the whole structure condensed. At this time, loops of FtsZ-GFP polymers emanated outward from the condensing ring structure and other unstable fluorescent structures elsewhere in the cell were also observed. Assembly of FtsZ rings at new division sites occurred within 1 min, from what appeared to be single points. Interestingly, this nucleation often took place in the predivisional cell at the same time the central FtsZ ring was in its final contraction phase. This demonstrates directly that, at least when FtsZ-GFP is being expressed, new division sites have the capacity to become fully functional for FtsZ targeting and assembly before cell division of the mother cell is completed. The results suggest that the timing of FtsZ assembly may be normally controlled in part by cellular FtsZ concentration. The use of wide-field optical sectioning microscopy to obtain sharp fluorescence images of FtsZ structures is also discussed.

Bacterial Proteins↗

PubMed1998

Effect of Pseudomonas fluorescens on beef discoloration and oxymyoglobin oxidation in vitro.

The relationship between bacterial growth and oxymyoglobin oxidation in vitro and in meat was studied. In the in vitro study, oxymyoglobin was combined with Pseudomonas fluorescens or sterile nutrient broth (control) in an airtight vessel. P. fluorescens samples showed greater metmyoglobin formation and oxygen consumption than controls. The P. fluorescens population in the reaction vessels was correlated with metmyoglobin formation (r = 0.85, P < 0.05) and oxygen consumption (r = 0.91, P < 0.05). When P. fluorescens and oxymyoglobin were combined in an airtight vessel, reducing the headspace from 13 ml and 9 ml to 3 ml resulted in greater metmyoglobin formation (P < 0.05). In the meat study, beef cores prepared from longissimus lumborum were inoculated with P. fluorescens (10(7) CFU/cm2) or sterile peptone water (control), packaged under 1% O2 (+99% N2), air, or 100% O2 and stored at 4 degrees C. Inoculated beef cores showed higher bacterial loads and metmyoglobin formation than their respective controls during 10 h storage in 1% O2, 3 days in air, and 7 days in 100% O2 (P < 0.05). This finding indicated that P. fluorescens could accelerate beef discoloration. Overall, studies demonstrated that oxygen consumption concomitant with P. fluorescens growth decreased partial oxygen pressure, which accelerated oxymyoglobin oxidation.

Animals↗

PubMed1998

Construction of human insulin gene expression recombinant and its effect on blood glucose of diabetic rats.

OBJECTIVE: To study the effect of human insulin gene modified fibroblasts on blood glucose in diabetic rats. METHODS: An expression vector containing human insulin gene was constructed by recombinant DNA technique and introduced into fibroblast Ltk- cells by lipofectin-mediated DNA transfection. Following G418 screening, the survived cells were selected and enriched. Finally a cell line, PRI-12 was generated for the highest insulin production. Then, these cells were injected into streptozocin (STZ)-induced diabetic rats. RESULTS: Insulin DNA transfected Ltk- cells were able to express insulin at a high level in a long-term culture. Furthermore, these Ltk- transfectants could decrease blood glucose significantly (P < 0.01) and obviously increase the body weight (P < 0.01) when they were injected into STZ-induced diabetic rats. CONCLUSIONS: These results suggest that the cell lines transfected insulin gene could secrete insulin and execute effect on diabetic rats. The study provides support for the view that somatic cell gene therapy offers a potential approach to delivery insulin into diabetes mellitus.

Animals↗

PubMed1998

[Chemical constituents of Quercus mongolica Fisch].

OBJECTIVE: To study the chemical constituents of the leaves of Quercus mongolica. METHOD: Using techniques of silical gel column chromatography and HNMR, 13CNMR. RESULT: Four compounds were isolated and identified as friedelin, glutinol, lupeo and beta-sitosterol respectively. CONCLUSION: All the compounds were isolated from this plant for the first time.

Drugs, Chinese Herbal↗

PubMed1998

[Identificating of Actinomycetes producting activity of herbicide].

14 strains of Actinomycetes were isolated from soil samples collected in Yunnan, China. Experiments showed that they had activity of herbicide. In this paper, the morphological and chemotaxonomic characters of these strains were studied and they were identified as three genera of Actinomycetales respecitively, such as Streptomyces, Micromonospora and Nocardia.

Actinobacteria↗

PubMed1998

[Effect of changes of amino acids of N-terminal region of the mature protein on secretion of alpha-amylase in B. subtilis].

The mutant plasmid pAmy413C, in which G takes the place of A at the 271 position of alpha-amylase gene on the pAmy413 from B. licheniformis, was constructed by site-direct mutagenesis. At the N-terminus of the mature alpha-amylase, amino acid +2Asn was substituted by +3Asp in the wild type protein. Then, the alpha-amylase output of the mutant plasmid pAmy413C in B. subtilis was 2.02-2.57 times higher than that of the wild type pAmy413C in the same strain. The amino acid sequencing at the N-terminus of the matural alpha-amylase revealsed that the recognition site of signal peptidase I moved one amino acid upstream, from Ala-(+2)Asn to AlaAla-(+3) Asp. That is, the +2Asn of the wild type was changed to the +3Asp of the mutant. The secondary structural analysis showed that a 14-cycle structure formed in the alpha-amylase mRNA when the free energy was -51.7 kcal. In this case, the mutant is identical with the wild type. The difference between them is that G at 271 position is no longer paired with U at 211 position, hence, a G-overhang is formed. The secondary structural analysis of protein showed that one amino acid diminished in the turn structure of amino acid at 33-37 position, and this very amino acid is involed in an alpha-helix structure. In short, all the changes mentioned above in conformation and charged amino acids contribute to the increase in the protein secretion in B. subtilis.

Amino Acid Sequence↗

PubMed1997

A terbenzimidazole that preferentially binds and conformationally alters structurally distinct DNA duplex domains: a potential mechanism for topoisomerase I poisoning.

The terbenzimidazoles are a class of synthetic ligands that poison the human topoisomerase I (TOP1) enzyme and promote cancer cell death. It has been proposed that drugs of this class act as TOP1 poisons by binding to the minor groove of the DNA substrate of TOP1 and altering its structure in a manner that results in enzyme-mediated DNA cleavage. To test this hypothesis, we characterize and compare the binding properties of a 5-phenylterbenzimidazole derivative (5PTB) to the d(GA4T4C)2 and d(GT4A4C)2 duplexes. The d(GA4T4C)2 duplex contains an uninterrupted 8-bp A.T domain, which, on the basis of x-ray crystallographic data, should induce a highly hydrated "A-tract" conformation. This duplex also exhibits anomalously slow migration in a polyacrylamide gel, a feature characteristic of a noncanonical global conformational state frequently described as "bent." By contrast, the d(GT4A4C)2 duplex contains two 4-bp A.T tracts separated by a TpA dinucleotide step, which should induce a less hydrated "B-like" conformation. This duplex also migrates normally in a polyacrylamide gel, a feature further characteristic of a global, canonical B-form duplex. Our data reveal that, at 20 degrees C, 5PTB exhibits an approximately 2. 3 kcal/mol greater affinity for the d(GA4T4C)2 duplex than for the d(GT4A4C)2 duplex. Significantly, we find this sequence/conformational binding specificity of 5PTB to be entropic in origin, an observation consistent with a greater degree of drug binding-induced dehydration of the more solvated d(GA4T4C)2 duplex. By contrast with the differential duplex affinity exhibited by 5PTB, netropsin and 4',6-diamidino-2-phenylindole (DAPI), two AT-specific minor groove binding ligands that are inactive as human TOP1 poisons, bind to both duplexes with similar affinities. The electrophoretic behaviors of the ligand-free and ligand-bound duplexes are consistent with 5PTB-induced bending and/or unwinding of both duplexes, which, for the d(GA4T4C)2 duplex, is synergistic with the endogenous sequence-directed electrophoretic properties of the ligand-free duplex state. By contrast, the binding to either duplex of netropsin or DAPI induces little or no change in the electrophoretic mobilities of the duplexes. Our results demonstrate that the TOP1 poison 5PTB binds differentially to and alters the structures of the two duplexes, in contrast to netropsin and DAPI, which bind with similar affinities to the two duplexes and do not significantly alter their structures. These results are consistent with a mechanism for TOP1 poisoning in which drugs such as 5PTB differentially target conformationally distinct DNA sites and induce structural changes that promote enzyme-mediated DNA cleavage.

Base Sequence↗

PubMed1997

Different HPV16 E6/E7 oncogene expression patterns in epithelia reconstructed from HPV16-immortalized human endocervical cells and genital keratinocytes.

Human papillomavirus type 16 (HPV16) E6/E7 oncogenes immortalize two types of human genital epithelial cells in vitro, endocervical cells and ectocervical or foreskin keratinocytes. Epithelia reconstructed in in vivo nude mouse implants or in vitro organotypic raft cultures from immortalized endocervical cells form higher grade dysplasia than those from keratinocytes. Here, we compared viral E6/E7 mRNA expression in immortalized cell lines of the three cell types using implants, rafts and in situ hybridization assays. Endocervical cells expressed E6/E7 throughout their reconstructed epithelia. In contrast, oncogenes were limited to basal cells for keratinocyte lower grade dysplasias. To study the role of the HPV16 promoter/enhancer in this repression in the upper layers of keratinocyte epithelia, new cell lines were established by immortalization with E6/E7 controlled by the SV40 promoter. The oncogenes were shown to be controlled from the SV40 elements after immortalization. Nevertheless, E6/E7 in the two cell types had the same cell-specific expression pattern as that controlled from the homologous HPV16 promoter. In addition, naturally occurring premalignant lesions having integrated HPV16 DNA expressed E6/E7 extensively in the high-grade dysplastic region of undifferentiated metaplasia. On the other hand, oncogene expression was restricted to lower layers in the lower grade dysplastic region of more mature differentiation. Our data suggest that keratinocytes have an inherent HPV16 promoter-nonspecific mechanism of repression. Apparently this mechanism, which can be acquired during maturation, is initially nonfunctional in in vitro and in vivo epithelia derived from metaplastic endocervical cells.

Animals↗

PubMed1997

Roles of the N and C terminal domains of the interleukin-3 receptor alpha chain in receptor function.

The interleukin-3 (IL-3), granulocyte-macrophage colony-stimulating factor, and IL-5 receptor alpha chains are each composed of three extracellular domains, a transmembrane domain and a short intracellular region. Domains 2 and 3 constitute the cytokine receptor module (CRM), typical of the cytokine receptor superfamily; however, the function of the N-terminal domain is not known. We have investigated the functions of the N-terminal and C-terminal domains of the IL-3 receptor (IL-3R) alpha chain. We find that cells transfected with the receptor beta chain (h beta c) and a truncated IL-3R alpha that is devoid of the intracellular region fail to proliferate or to activate STAT5 in response to human IL-3, despite binding the IL-3 with affinity indistinguishable from that of full-length receptor. In addition, IL-3-induced phosphorylation of h beta c was not detected. Thus, the IL-3R alpha intracellular region does not contribute detectably to stabilization of the receptor/ligand complex, but is essential for signal propagation. In contrast, a truncated IL-3R alpha with the N-terminal domain deleted interacts functionally with the beta chain; mouse cells transfected with these receptor chains proliferate in response to human IL-3 and STAT5 transcription factor is activated. High- and low-affinity binding sites are retained, although the affinity for IL-3 is decreased 15-fold, indicating a significant role for the N-terminal domain in IL-3 binding.

Amino Acid Sequence↗

PubMed1997

Tyrosine phosphorylation and axon guidance: of mice and flies.

Recent genetic evidence suggests that tyrosine kinases and tyrosine phosphatases can control the guidance of specific growth cones. Within a family of related phosphatases or kinases, individual members can have partially redundant functions. Receptor phosphatases can work together at one guidance choice point, but in opposition at another. The specific combination of kinases and phosphatases active in a growth cone may be an important determinant of pathway choice. One mechanism by which these proteins could control guidance decisions is through regulation of adhesion between growth cones and axons.

Animals↗