Comparisons of two methods for haplotype reconstruction and haplotype frequency estimation from population data.
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Algorithms↗
Biomedical subjects
S Zhang
Publications and source records attributed to S Zhang.
Immunolocalization of odorant-binding proteins in noctuid moths (Insecta, Lepidoptera).
Odorant-binding proteins were studied in the noctuid moths Agrotis segetum, Autographa gamma, Helicoverpa armigera, Heliothis virescens and Spodoptera littoralis using antisera raised against the pheromone-binding protein (PBP) and general odorant-binding protein 2 (GOBP2) of Antheraea polyphemus (Saturniidae). Proteins immunoreacting with these antisera were only found on the antennae and PBP and GOBP2 could be identified on western blots of males and females of all five species. PBPs were predominantly localized in sensilla trichodea and GOBP2 in sensilla basiconica, in good correlation with the stimulus specificity of the receptor cells in these sensilla. In H. armigera and H. virescens the majority of the s. trichodea immunoreacted with the antiserum against PBP of A. polyphemus; in A. segetum, A. gamma and S. littoralis, on the other hand, a high percentage of s. trichodea remained unlabelled. Probably, the PBP expressed in these sensilla is so different that it does not immunoreact with the antiserum used. Such a protein was found by native PAGE of antennal extracts of A. segetum and S. littoralis. These data correlate with the fact that the two heliothine species use pheromones with the same alkyl chain length as A. polyphemus, while the other three species use pheromones with shorter chains. In H. armigera, H. virescens, A. gamma and S. littoralis female antennae were also immunolabelled and a large number of PBP-expressing s. trichodea was consistently found. In S.littoralis this fits with the electrophysiologically recorded high pheromone sensitivity of female s. trichodea, whereas in females of H. armigera and H. virescens no or only weak responses to pheromone stimulation have been reported. Therefore, PBP expression in a sensillum does not necessarily imply pheromone sensitivity of its receptor cells.
Animals↗
Previous pulmonary diseases and risk of lung cancer in Gansu Province, China.
BACKGROUND: Although active smoking is well established as the main cause of lung cancer, there is accumulating evidence that history of prior lung diseases may be an independent risk factor for lung cancer. METHODS: A population-based case-control study in Gansu Province, China identified 886 lung cancer cases (656 male, 230 female) diagnosed between January 1994 and April 1998. A standardized interview collected information on a variety of potential risk factors including a history of physician-diagnosed non-malignant lung diseases (pulmonary tuberculosis, chronic bronchitis/emphysema, asthma, pneumonia), age and year in which each condition was first diagnosed, and any therapy or hospitalization received. RESULTS: Pulmonary tuberculosis (odds ratio [OR] = 2.1, 95% CI : 1.4-3.1) and chronic bronchitis/emphysema (OR = 1.4, 95% CI : 1.1-1.8) were associated with increased risk of lung cancer, after adjustment for active smoking and socioeconomic status. The OR for asthma (OR = 1.4, 95% CI : 0.9-2.1) and pneumonia (OR = 1.5, 95% CI : 1.0-2.3) were also elevated. The risk of lung cancer remained significant for pulmonary tuberculosis and chronic bronchitis/emphysema when analysis was limited to the pathologically confirmed cases and self-responders. CONCLUSIONS: This study provides additional evidence that previous pulmonary tuberculosis and chronic bronchitis/emphysema are causally related to lung cancer, although the precise mechanism is still unclear. The results for asthma and pneumonia, while suggestive of a positive association, did not reach the traditional level of statistical significance and should be interpreted with caution.
Adult↗
Carriage of antibiotic-resistant bacteria by healthy children.
The frequency of carriage of antibiotic-resistant bacteria in healthy 7- and 8-year-old children in Bristol was studied. Children born in Avon between 1 April 1991 and 31 December 1992, attending the Avon Longitudinal Study of Pregnancy and Childhood (ALSPAC) 7 year follow-up clinic, formed the study population. Carriage was estimated using mouth and stool samples. None of 105 children on whom information was available had received tetracycline, chloramphenicol, ciprofloxacin or an extended-spectrum cephalosporin in the previous year. Staphylococcus aureus was isolated from mouthwashes from 200 (37.1%) of 539 children sampled. Six (3%) of the isolates were resistant to chloramphenicol or tetracycline and four (2%) were methicillin resistant. Haemophilus spp. were isolated from 369 (72%) of 513 samples and 63 (17%) were ampicillin resistant, 49 (13.3%) were erythromycin resistant and seven (1.9%) were tetracycline resistant. Branhamella catarrhalis was isolated from 333 (74%) of 450 samples. Twenty-eight (8.4%) were erythromycin resistant and 14 (4.2%) strains were tetracycline resistant. Group A beta-haemolytic streptococci were isolated from 17 of 507 children sampled. One (5.9%) was tetracycline resistant. Stool samples were returned from 335 (62%) of 539 children from whom they were requested. Eleven per cent of samples yielded Gram-negative bacilli with high-level resistance to chloramphenicol, which was frequently linked to resistance to ampicillin, spectinomycin and streptomycin. Isolates demonstrating resistance to the third-generation cephalosporin ceftazidime were recovered from 17 subjects (3.2%). Six (35%) of 17 isolates possessed extended-spectrum beta-lactamases. Healthy children carry bacteria resistant to antibiotics to which children are not usually exposed. Resistance to ceftazidime, chloramphenicol and tetracycline may be co-selected by exposure to other antibiotics used in children or may be acquired from family members, pets, other children or food. These results suggest that antibiotic-resistant bacteria are widely disseminated and may be acquired by children before exposure to specific selection pressure.
Bacteria↗
Only the reduced conformer of alpha-lactalbumin is inducible to aggregation by protein aggregates.
Reduced apo-alpha-lactalbumin (r-LA) in the pre-molten globule state is soluble in neutral and reduced buffer at 25 degrees C but becomes aggregated when aggregates of various proteins are added. However, protein aggregates do not induce the aggregation of apo-alpha-lactalbumin in the molten globule state. The presence of the molecular chaperone protein disulfide isomerase or the "chemical chaperone" polyethyleneglycol inhibits the induced aggregation. Native proteins, aggregation-free folding intermediates, and soluble aggregates do not induce the aggregation. The interaction between r-LA and protein aggregates is hydrophobic in nature. These findings suggest that pre-molten globule state of LA is the target not only for chaperones but also for protein aggregates.
Animals↗
Extracellular calmodulin stimulates light-independent RbcS-GUS expression in suspension-cultured cells of transgenic tobacco.
The RbcS genes encode the small subunits of rubisco; the expression of these genes is controlled in a light-dependent and independent manner. It has been reported that intracellular calmodulin (CaM) is involved in light-dependent RbcS expression. In this report, the role of extracellular CaM in regulating expression of RbcS in darkness was examined. The time course of expression of RbcS-GUS and that of the secretion of CaM in the suspended transgenic tobacco cells in darkness were very similar. Both showed initial increase followed by decline with maximum CaM secretion preceding maximum GUS expression by 24 h. The concentration of CaM in the culture medium is regulated light independently. Purified CaM alone added to the media enhanced RbcS-GUS expression in darkness. The addition of membrane-impermeable CaM inhibitors, such as anti-CaM antiserum or W7-agarose, repressed the expression of RbcS-GUS in darkness, but this inhibitory effect was completely reversed by adding exogenous purified CaM. These results provide the first evidence that extracellular CaM is involved in the regulation of light-independent RbcS gene expression.
Calmodulin↗
Molecular genetic evidence for different clonal origin of components of human renal angiomyolipomas.
Renal angiomyolipoma is a benign neoplasm composed of variable proportions of blood vessels, smooth muscle, and adipose tissue. Smooth muscle, adipose tissue, blood vessels, and adjacent normal kidney tissue were separately microdissected from sections prepared from formalin-fixed, paraffin-processed tissues from angiomyolipomas from 18 women. X chromosome inactivation analysis using the methylation pattern at exon 1 of the human androgen receptor gene on chromosome Xq11-12 was used to study the clonal origin of each component. Nonrandom inactivation of X chromosomes was found in six of the 15 informative tumors. The smooth muscle and adipose tissue showed differing patterns of nonrandom inactivation of X chromosomes in five angiomyolipomas and the same pattern of nonrandom inactivation of X chromosomes in one. Samples from the blood vessels showed random inactivation of X chromosomes in all informative cases. Our data showed that the adipose tissue and smooth muscle cells of renal angiomyolipoma are both monoclonal but may arise independently. The coexistence of tumor subclones with morphologic heterogeneity can lead to the formation of a clinically detectable tumor.
Adipose Tissue↗
Gd3+ complexes with slowly exchanging bound-water molecules may offer advantages in the design of responsive MR agents.
RATIONALE AND OBJECTIVES: Slow water exchange in Gd3+ complexes is generally considered detrimental to their use as MR contrast agents. The objective of this work was to demonstrate how this feature may serve as a useful template for the design of responsive MR agents. METHODS: Lanthanide (Ln) complexes of two 1,4,7,10-tetraazacyclododecane-N,N',N",N'"-tetraacetic acid (DOTA)-tetraamide phosphonate (1) and phosphonate ester (2) ligands were studied by multinuclear (1H, 13C, 31P, and 17O) nuclear MR spectroscopy. RESULTS: The inner-sphere water lifetime in the Ln(2) complexes was much longer (tauM298 = 0.8-1.3 ms) than in the corresponding Ln(1) complexes. This allowed direct detection of the bound-water molecule in europium(2) in water at 40 degrees C by 1H nuclear MR. The water relaxivity of gadolinium(2) was independent of pH between 8.5 and 6.0, whereas the relaxivity of gadolinium(1) increased more than twofold in this pH range. CONCLUSIONS: T1-weighted images of phantoms containing gadolinium(1) at different pH values demonstrate the efficacy of this complex as a pH-sensitive MR contrast agent.
Contrast Media↗
Gene expression profiling in human fetal liver and identification of tissue- and developmental-stage-specific genes through compiled expression profiles and efficient cloning of full-length cDNAs.
Fetal liver intriguingly consists of hepatic parenchymal cells and hematopoietic stem/progenitor cells. Human fetal liver aged 22 wk of gestation (HFL22w) corresponds to the turning point between immigration and emigration of the hematopoietic system. To gain further molecular insight into its developmental and functional characteristics, HFL22w was studied by generating expressed sequence tags (ESTs) and by analyzing the compiled expression profiles of liver at different developmental stages. A total of 13,077 ESTs were sequenced from a 3'-directed cDNA library of HFL22w, and classified as follows: 5819 (44.5%) matched to known genes; 5460 (41.8%) exhibited no significant homology to known genes; and the remaining 1798 (13.7%) were genomic sequences of unknown function, mitochondrial genomic sequences, or repetitive sequences. Integration of ESTs of known human genes generated a profile including 1660 genes that could be divided into 15 gene categories according to their functions. Genes related to general housekeeping, ESTs associated with hematopoiesis, and liver-specific genes were highly expressed. Genes for signal transduction and those associated with diseases, abnormalities, or transcription regulation were also noticeably active. By comparing the expression profiles, we identified six gene groups that were associated with different developmental stages of human fetal liver, tumorigenesis, different physiological functions of Itoh cells against the other types of hepatic cells, and fetal hematopoiesis. The gene expression profile therefore reflected the unique functional characteristics of HFL22w remarkably. Meanwhile, 110 full-length cDNAs of novel genes were cloned and sequenced. These novel genes might contribute to our understanding of the unique functional characteristics of the human fetal liver at 22 wk.
Cloning, Molecular↗
Activation of salicylic acid-induced protein kinase, a mitogen-activated protein kinase, induces multiple defense responses in tobacco.
The activation of mitogen-activated protein kinases (MAPKs) is one of the earliest responses in plants challenged by avirulent pathogens or cells treated with pathogen-derived elicitors. Expression of a constitutively active MAPK kinase, NtMEK2(DD), in tobacco induces the expression of defense genes and hypersensitive response-like cell death, which are preceded by the activation of two endogenous MAPKs, salicylic acid-induced protein kinase (SIPK) and wounding-induced protein kinase (WIPK). However, the roles that SIPK and WIPK each play in the process are unknown. Here we report that SIPK alone is sufficient to activate these defense responses. In tobacco leaves transiently transformed with SIPK under the control of a steroid-inducible promoter, the induction of SIPK expression after the application of dexamethasone, a steroid, leads to an increase of SIPK activity. The increase of SIPK activity is dependent on the phosphorylation of newly synthesized SIPK by its endogenous upstream kinase. In contrast, the expression of WIPK under the same conditions fails to increase its activity, even though the protein accumulates to a similar level. Studies using chimeras of SIPK and WIPK demonstrated that the C terminus of SIPK contains the molecular determinant for its activation, which is rather surprising because the N termini of SIPK and WIPK are more divergent. SIPK has been implicated previously in the regulation of both plant defense gene activation and hypersensitive response-like cell death based on evidence from pharmacological studies using kinase inhibitors. This gain-of-function study provided more direct evidence for its role in the signaling of multiple defense responses in tobacco.
Amino Acid Sequence↗
Cocaine blocks HERG, but not KvLQT1+minK, potassium channels.
Cocaine causes cardiac arrhythmias, sudden death, and occasionally long QT syndrome in humans. We investigated the effect of cocaine on the human K(+) channels HERG and KvLQT1+minK that encode native rapidly (I(Kr)) and slowly (I(Ks)) activating delayed rectifier K(+) channels in the heart. HERG and KvLQT1+minK channels were heterologously expressed in human embryonic kidney 293 cells, and whole-cell currents were recorded. Cocaine had no effect on KvLQT1+minK current in concentrations up to 200 microM. In contrast, cocaine reversibly blocked HERG current with half-maximal block of peak tail current of 7.2 microM. By using a protocol to quickly activate HERG channels, we found that cocaine block developed rapidly after channel activation. At 0 mV, the time constants for the development of block were 38.2 +/- 2.1, 15.2 +/- 0.8, and 6.9 +/- 1.1 ms in 10, 50 and 200 microM cocaine, respectively. Cocaine-blocked channels also recovered rapidly from block after repolarization. At -100 mV, recovery from block followed a biphasic time course with fast and slow time constants of 3.5 +/- 0.7 and 100.3 +/- 15.4 ms, respectively. Using N-methyl-cocaine, a permanently charged, membrane-impermeable cocaine analog, block of HERG channels rapidly developed when the drug was applied intracellularly through the patch pipette, suggesting that the cocaine binding site on the HERG protein is located on a cytoplasmic accessible domain. These results indicate that cocaine suppresses HERG, but not KvLQT1+minK, channels by preferentially blocking activated channels, that it unblocks upon repolarization, and does so with unique ultrarapid kinetics. Because the cocaine concentration range we studied is achieved in humans, HERG block may provide an additional mechanism for cocaine-induced arrhythmias and sudden death.
Cation Transport Proteins↗
Toxic effects of Ag(I) and Hg(II) on Candida albicans and C. maltosa: a flow cytometric evaluation.
The effects of Ag(I) and Hg(II) on membrane potential and integrity of cells of Candida albicans and C. maltosa were determined with a flow cytometric procedure that employed an anionic membrane potential-sensitive dye, bis-(1,3-dibutylbarbituric acid) trimethine oxonol, and a membrane integrity indicator, propidium iodide. The membrane potentials of cells of both species were reduced rapidly within 15 min of exposure to Ag(I). No threshold dose for Hg(II) existed, and cells of both species lost membrane potential gradually in Hg(II) solutions. Cells of both species lost membrane integrity more rapidly in Ag(I) solutions than in Hg(II) solutions. In Ag(I) solutions, the decrease in the numbers of cells recoverable in culture occurred at a rate similar to the rate of cell depolarization and membrane permeabilization. In Hg(II) solutions, loss of cell recoverability preceded the loss of membrane potential and membrane integrity. C. albicans, in contrast to C. maltosa, showed no loss of membrane integrity after exposure to Hg(II) solutions for 1 h. Different rates of binding of Ag(I) and Hg(II) between the two species suggest that the two ions target different primary sites.
Candida↗
Differentiation of two bovine lentiviruses by a monoclonal antibody on the basis of epitope specificity.
Bovine immunodeficiency virus (BIV) and Jembrana disease virus (JDV) are bovine lentiviruses that are closely related genetically. A recombinant fusion protein containing the capsid protein of BIV expressed in Escherichia coli was used to immunize mice and produce monoclonal antibodies. Six hybridomas specific for BIV capsid protein were identified, and one antibody, designated 10H1, was characterized further. Competitive binding assays were performed to analyze the topography of antigenic determinants by enzyme-linked immunosorbent assay and demonstrated the existence of at least three distinct antigenic determinants on capsid protein. The monoclonal antibody reacted specifically with both BIV capsid and the recombinant fusion protein in Western immunoblot analyses. However, it did not react with the recombinant capsid fusion protein of JDV, indicating that BIV contains at least one unique epitope in the capsid protein that is absent in JDV. Further mapping of the epitope by chemical cleavage analysis identified that the epitope is located at the 6.4-kDa N terminus of the 29-kDa capsid protein. This monoclonal antibody assay will be valuable for distinguishing the two closely related lentiviruses by Western blotting.
Animals↗
Salmonella-induced cell death is not required for enteritis in calves.
Salmonella enterica serovar Typhimurium causes cell death in bovine monocyte-derived and murine macrophages in vitro by a sipB-dependent mechanism. During this process, SipB binds and activates caspase-1, which in turn activates the proinflammatory cytokine interleukin-1beta through cleavage. We used bovine ileal ligated loops to address the role of serovar Typhimurium-induced cell death in induction of fluid accumulation and inflammation in this diarrhea model. Twelve perinatal calves had 6- to 9-cm loops prepared in the terminal ileum. They were divided into three groups: one group received an intralumen injection of Luria-Bertani broth as a control in 12 loops. The other two groups (four calves each) were inoculated with 0.75 x 10(9) CFU of either wild-type serovar Typhimurium (strain IR715) or a sopB mutant per loop in 12 loops. Hematoxylin and eosin-stained sections were scored for inflammation, and terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling (TUNEL)-positive cells were detected in situ. Fluid accumulation began at 3 h postinfection (PI). Inflammation was detected in all infected loops at 1 h PI. The area of TUNEL-labeled cells in the wild-type infected loops was significantly higher than that of the controls at 12 h PI, when a severe inflammatory response and tissue damage had already developed. The sopB mutant induced the same amount of TUNEL-positive cells as the wild type, but it was attenuated for induction of fluid secretion and inflammation. Our results indicate that serovar Typhimurium-induced cell death is not required to trigger an early inflammatory response and fluid accumulation in the ileum.
Animals↗
RNA expression analysis using an antisense Bacillus subtilis genome array.
We have developed an antisense oligonucleotide microarray for the study of gene expression and regulation in Bacillus subtilis by using Affymetrix technology. Quality control tests of the B. subtilis GeneChip were performed to ascertain the quality of the array. These tests included optimization of the labeling and hybridization conditions, determination of the linear dynamic range of gene expression levels, and assessment of differential gene expression patterns of known vitamin biosynthetic genes. In minimal medium, we detected transcripts for approximately 70% of the known open reading frames (ORFs). In addition, we were able to monitor the transcript level of known biosynthetic genes regulated by riboflavin, biotin, or thiamine. Moreover, novel transcripts were also detected within intergenic regions and on the opposite coding strand of known ORFs. Several of these novel transcripts were subsequently correlated to new coding regions.
Amino Acid Sequence↗
Loss of ribosomal protein L11 blocks stress activation of the Bacillus subtilis transcription factor sigma(B).
sigma(B), the general stress response sigma factor of Bacillus subtilis, is activated when the cell's energy levels decline or the bacterium is exposed to environmental stress (e.g., heat shock, ethanol). Physical stress activates sigma(B) through a collection of regulatory kinases and phosphatases (the Rsb proteins) which catalyze the release of sigma(B) from an anti-sigma(B) factor inhibitor. The means by which diverse stresses communicate with the Rsb proteins is unknown; however, a role for the ribosome in this process was suggested when several of the upstream members of the sigma(B) stress activation cascade (RsbR, -S, and -T) were found to cofractionate with ribosomes in crude B. subtilis extracts. We now present evidence for the involvement of a ribosome-mediated process in the stress activation of sigma(B). B. subtilis strains resistant to the antibiotic thiostrepton, due to the loss of ribosomal protein L11 (RplK), were found to be blocked in the stress activation of sigma(B). Neither the energy-responsive activation of sigma(B) nor stress-dependent chaperone gene induction (a sigma(B)-independent stress response) was inhibited by the loss of L11. The Rsb proteins required for stress activation of sigma(B) are shown to be active in the RplK(-) strain but fail to be triggered by stress. The data demonstrate that the B. subtilis ribosomes provide an essential input for the stress activation of sigma(B) and suggest that the ribosomes may themselves be the sensors for stress in this system.
Bacillus subtilis↗
Kilham polyomavirus: activation of gene expression and DNA replication in mouse fibroblast cells by an enhancer substitution.
The Kilham strain of polyomavirus (KV) infects vascular endothelial cells in vivo (J. E. Greenlee, Infect. Immun. 26:705-713, 1979), but no permissive cell type for growth of the virus in vitro has been identified. The failure of KV DNA to replicate in mouse fibroblast cells after transfection suggested that viral gene expression had narrow cell specificity. A KV substitution mutant having a part of the regulatory region of KV DNA replaced with a segment of the polyomavirus transcriptional enhancer was constructed. The substitution mutant was able to replicate in transfected 3T3 cells, and the newly replicated viral DNA associated with protein to form particles with the density of virions in CsCl equilibrium gradients. However, these particles were noninfectious when tested on 3T3 cells, suggesting that absorption or uptake of virus particles was defective for these cells. Analysis of early and late promoter activities by luciferase reporter gene expression showed that the enhancer substitution had a moderate positive effect on early gene expression and a large effect on the expression of the late genes. KV large T antigen inhibited the activities of both the wild-type and the substitution mutant early promoter, whereas only the mutant late promoter was activated under the same conditions. A comparison of the KV and polyomavirus large T antigens showed that they were not interchangeable in the initiation of KV and polyomavirus DNA synthesis. Furthermore, the wild-type KV origin of DNA replication was less active than the mutant structure in the presence of saturating amounts of KV large T antigen. Together, our data demonstrate several differences between the two types of large T antigen in their interactions with cellular proteins.
Animals↗
Urinary 1-hydroxypyrene in coke oven workers relative to exposure, alcohol consumption, and metabolic enzymes.
OBJECTIVES: To investigate the influence of personal lifestyle--such as smoking and alcohol consumption-on urinary 1-hydroxypyrene (1-OHP) concentrations in coke oven workers exposed to polycyclic aromatic hydrocarbons (PAHs) and to evaluate the association of 1-OHP concentrations with the genetic polymorphism of several metabolic enzymes including cytochrome P-450 (CYP) 1A1 and glutathione S-tranferases (GSTs). METHODS: The study population contained 162 coke oven workers and 58 controls employed at the largest iron and steel factory in China. Personal data were collected at the interview. 1-OHP in urine was measured with high performance liquid chromatography with fluorescence detection. Genetic polymorphisms were identified by the polymerase chain reaction (PCR) method. RESULTS: A positive association between excretion of urinary 1-OHP and the levels of exposure to PAHs was confirmed. Those people who consumed >or=50 g/day ethanol had significantly higher 1-OHP excretion than did other coke oven workers (p<0.01). No significant difference in urinary 1-OHP was found between smokers and non-smokers, in both controls and exposed subjects. The variant homozygotes at exon 7 of the CYP1A1 gene had significantly higher urinary 1-OHP concentrations than other CYP1A1 genotypes among the exposed workers (p=0.03). There was less association between the concentrations of 1-OHP and the GSTM1, GSTP1, or GSTT1 polymorphism. CONCLUSIONS: The present study confirmed that urinary 1-OHP is a good biomarker for exposure to PAHs. Alcohol consumption affected urinary 1-OHP excretion. The variant genotypes of the CYP1A1 gene may result in the enhancement of PAH metabolites. It is helpful to understand the role of individual susceptibility on metabolism of carcinogens. These findings suggest that the modulating effect of individual lifestyle factors or genetic nature should be considered in future studies on occupational exposure to PAHs and in evaluating the health risk from harmful chemicals.
Adult↗