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David R Nelson

Publications and source records attributed to David R Nelson.

At least 73 records · Page 4Linked to original sources
PubMed2004

Regulation of Vibrio anguillarum empA metalloprotease expression and its role in virulence.

Atlantic salmon (Salmo salar) were challenged with Vibrio anguillarum strains M93Sm and NB10 and empA null mutants M99 and NB12. Both wild types were virulent when administered by intraperitoneal (i.p.) injection or anal intubation. NB12 was avirulent via either route of infection. M99 virulence was attenuated when delivered by intubation, but fully virulent by i.p. injection. Northern blot analysis revealed empA expression in M93Sm and NB10 cells incubated in mucus, while incubation in Luria-Bertani broth plus 2% NaCl (LB20) induced empA expression only in NB10. Nucleotide differences between M93Sm and NB10 empA sequences were found in regions located 207 and 229 bp upstream of the empA translational start. Reverse transcription-PCR and 5' rapid amplification of cDNA ends revealed the empA transcriptional start site 85 bp upstream of the translational start for both strains. A putative sigma(S)-dependent promoter was identified upstream of the transcriptional start in both strains. Site-directed mutagenesis was used to create rpoS mutants of M93Sm and NB10. Neither rpoS mutant exhibited protease activity. Since empA is expressed during stationary phase, the effects of conditioned medium on protease activity were examined. M99 conditioned LB20 supernatants stimulated protease activity in NB10 while allowing M93Sm to produce protease in LB20. Neither acyl homoserine lactones nor AI-2 induced protease activity. Conditioned LB20 supernatant from a V. anguillarum luxS mutant caused a more rapid induction of protease activity in wild-type cells. Our data show that expression of empA is differentially regulated in V. anguillarum strains NB10 and M93Sm and requires sigma(S), quorum-sensing molecules, and gastrointestinal mucus.

Bacterial Proteins↗
PubMed2004

The truth about mouse, human, worms and yeast.

Genome comparisons are behind the powerful new annotation methods being developed to find all human genes, as well as genes from other genomes. Genomes are now frequently being studied in pairs to provide cross-comparison datasets. This 'Noah's Ark' approach often reveals unsuspected genes and may support the deletion of false-positive predictions. Joining mouse and human as the cross-comparison dataset for the first two mammals are: two Drosophila species, D. melanogaster and D. pseudoobscura; two sea squirts, Ciona intestinalis and Ciona savignyi; four yeast (Saccharomyces) species; two nematodes, Caenorhabditis elegans and Caenorhabditis briggsae; and two pufferfish (Takefugu rubripes and Tetraodon nigroviridis). Even genomes like yeast and C. elegans, which have been known for more than five years, are now being significantly improved. Methods developed for yeast or nematodes will now be applied to mouse and human, and soon to additional mammals such as rat and dog, to identify all the mammalian protein-coding genes. Current large disparities between human Unigene predictions (127,835 genes) and gene-scanning methods (45,000 genes) still need to be resolved. This will be the challenge during the next few years.

Algorithms↗
PubMed2004

'Frankenstein genes', or the Mad Magazine version of the human pseudogenome.

Annotation of the human genome is inching forward. Seven human chromosomes have now been fully annotated, covering 17 per cent of the genome, and at least one chromosome has been re-annotated. The enormity of the task forces a dependence on automated tools for detecting and assembling the genes, followed by hand curation to correct errors and polish the gene models. The accuracy of gene prediction algorithms is very good for internal exons from intact genes, but these programs do peculiar and exasperating things to pseudogenes. These programs can actually resurrect pseudogenes from the dead, making them into viable gene models for intact proteins, albeit science-fictional proteins. This process is demonstrated for four human pseudogenes from the cytochrome P450 family and one putatively functional P450 gene, CYP2U1, having a non-consensus intron boundary. These examples are offered as a call-to-arms to improve pseudogene prediction as an art in itself, and not as a by-product of gene annotation. Failure to do so will flood the databases with thousands of false-positive predictions. Indeed, they are already there.

Amino Acid Sequence↗
PubMed2004

Cyclophilin nomenclature problems, or, 'a visit from the sequence police'.

Why is agreement on one particular name for each gene important? As one genome after another becomes sequenced, it is imperative to consider the complexity of genes, genetic architecture, gene expression, gene-gene and gene-product interactions and evolutionary relatedness across species. To agree on a particular gene name not only makes one's own research easier, it aids automated text-mining algorithms and search engines, which are increasingly employed to find relationships in the millions of abstracts in the medical research literature and sequence databases. A common nomenclature system will also be helpful to the present generation, as well as future generations, of graduate students and postdoctoral fellows who are about to enter genomics research. In this paper, the authors present some problems that arose when two separate research communities decided to choose the same root, CYP, for naming their gene families. They then offer a logical solution, by renaming the cyclophilin genes with a common root, such as cyn- in Caenorhabditis and CYN- in mammals (Cyn in mouse), and using evolutionary divergence to cluster genes of the highest level of relatedness.

Amino Acid Sequence↗
PubMed2004

Predictors of long-term mortality in patients with cirrhosis of the liver admitted to a medical ICU.

CONTEXT: The long-term survival of patients with cirrhosis of the liver admitted to the ICU has not been described. OBJECTIVE: The main objectives were to determine the 1-year and 5-year mortality rates of a cohort of patients admitted to a medical ICU (MICU), and to identify the risk factors that may predict long-term outcomes. DESIGN: This is a cohort study. We used a model-building (MB) and model validation (MV) procedure that has previously been described to determine the risk factors for overall mortality. SETTINGS: An MICU in a major referral medical center. PATIENTS: Four hundred twenty consecutive patients admitted to the ICU from January 1, 1993, through October 31, 1998, met the inclusion criteria of diagnosis of liver failure, cirrhosis, chronic liver disease, variceal bleeding, hepatic encephalopathy, or hepatorenal syndrome. Patients with acute liver failure who had undergone liver transplantation, or candidates for orthotopic liver transplantation were excluded. INTERVENTION: None. RESULTS: The 1-year mortality rate was 69%, and the 5-year mortality rate was 77%. The median survival time was 1 month. The independent predictors of mortality in patients in the MB group who retained their significance in the MV group were as follows: an acute physiology, age, and chronic health evaluation (APACHE) III score of >/= 90 (hazard ratio [HR], 2.2; 95% confidence interval [CI], 1.6 to 2.8; p < 0.0001), the use of pressors (HR, 2.5; 95% CI, 1.9 to 3.2; p < 0.0001), and jaundice (HR, 1.7; 95% CI, 1.4 to 2.2; p < 0.0001). Patients with all three risk factors (ie, APACHE III score >/= 90, use of pressors, and jaundice) had a 92% 1-month mortality rate compared to 11.2% for patients with no risk factors. CONCLUSIONS: Patients admitted to an MICU with underlying cirrhosis who are not eligible for liver transplantation have a poor long-term prognosis, even if they survive the ICU admission, particularly as the number of risk factors increases.

APACHE↗
PubMed2004

Dynamics of molecular motors and polymer translocation with sequence heterogeneity.

The effect of sequence heterogeneity on polynucleotide translocation across a pore and on simple models of molecular motors such as helicases, DNA polymerase/exonuclease, and RNA polymerase is studied in detail. Pore translocation of RNA or DNA is biased due to the different chemical environments on the two sides of the membrane, whereas the molecular motor motion is biased through a coupling to chemical energy. An externally applied force can oppose these biases. For both systems we solve lattice models exactly both with and without disorder. The models incorporate explicitly the coupling to the different chemical environments for polymer translocation and the coupling to the chemical energy (as well as nucleotide pairing energies) for molecular motors. Using the exact solutions and general arguments, we show that the heterogeneity leads to anomalous dynamics. Most notably, over a range of forces around the stall force (or stall tension for DNA polymerase/exonuclease systems) the displacement grows sublinearly as t(micro), with micro < 1. The range over which this behavior can be observed experimentally is estimated for several systems and argued to be detectable for appropriate forces and buffers. Similar sequence heterogeneity effects may arise in the packing of viral DNA.

Algorithms↗
PubMed2003

Virus shapes and buckling transitions in spherical shells.

We show that the icosahedral packings of protein capsomeres proposed by Caspar and Klug for spherical viruses become unstable to faceting for sufficiently large virus size, in analogy with the buckling instability of disclinations in two-dimensional crystals. Our model, based on the nonlinear physics of thin elastic shells, produces excellent one-parameter fits in real space to the full three-dimensional shape of large spherical viruses. The faceted shape depends only on the dimensionless Foppl-von Kármán number gamma=YR(2)/kappa, where Y is the two-dimensional Young's modulus of the protein shell, kappa is its bending rigidity, and R is the mean virus radius. The shape can be parametrized more quantitatively in terms of a spherical harmonic expansion. We also investigate elastic shell theory for extremely large gamma, 10(3)<gamma<10(8), and find results applicable to icosahedral shapes of large vesicles studied with freeze fracture and electron microscopy.

Crystallography, X-Ray↗
PubMed2003

Cytochrome p450 complement (CYPome) of the avermectin-producer Streptomyces avermitilis and comparison to that of Streptomyces coelicolor A3(2).

The genus Streptomyces produces about two-thirds of naturally occurring antibiotics and a wide array of other secondary metabolites, including antihelminthic agents, antitumor agents, antifungal agents, and herbicides. The newly completed genome sequence of the avermectin-producing bacterium Streptomyces avermitilis contains 33 cytochromes p450 (CYPs), many more than the 18 observed in Streptomyces coelicolor A3(2). Some of the likely metabolic functions are reported together with their genomic location and bioinformatic analysis. Seven entirely new CYP families were found together with close homologues of some forms observed in S. coelicolor A3(2). The presence of unusual CYP forms associated with conservons is revealed and of these, CYP157 forms in both S. avermitilis and S. coelicolor A3(2) deviate from the previously accepted rule for an EXXR motif within the K-helix of CYPs. Amongst this range of CYPs are forms associated with avermectin, filipin, geosmin, and pentalenolactone biosynthesis as well as unknown pathways of secondary metabolism.

Chromosomes, Fungal↗
PubMed2003

Systemic host responses in severe sepsis analyzed by causative microorganism and treatment effects of drotrecogin alfa (activated).

Clinical trials with novel therapeutic agents for severe sepsis have suggested that patients might respond differently depending on causative microorganism. Data from a large, placebo-controlled trial of recombinant human drotrecogin alfa (activated) (DrotAA) were analyzed by type of causative microorganism for treatment-associated differences in mortality, coagulopathy, and inflammatory response. Compared with placebo, mortality rates associated with DrotAA were consistently reduced for each microorganism group (gram-positive bacteria, gram-negative bacteria, mixed bacteria, fungi, other, and unknown microbial etiology), with a stratified relative risk (RR) of 0.80 (95% confidence interval [CI], 0.69-0.94). The greatest reduction in the mortality rate was for Streptococcus pneumoniae infection (RR, 0.56; 95% CI, 0.35-0.88). Levels of coagulation and inflammation biomarkers varied with different pathogens at study entry. Results demonstrate that DrotAA, administered as an adjunct to standard anti-infective therapy, can improve the rate of survival for patients who develop severe sepsis regardless of causative microorganism.

Anti-Bacterial Agents↗
PubMed2003

Hexatic undulations in curved geometries.

We discuss the influence of two-dimensional hexatic order on capillary waves and undulation modes in spherical and cylindrical geometries. In planar geometries, extended bond-orientational order has only a minor effect on the fluctuations of liquid surfaces or lipid bilayers. However, in curved geometries, the long-wavelength spectrum of these ripples is altered. We calculate this frequency shift and discuss applications to spherical vesicles, liquid metal droplets, bubbles and cylindrical jets coated with surface-active molecules, and to multielectron bubbles in liquid helium at low temperatures. Hexatic order also leads to a shift in the threshold for the fission instability of charged droplets and bubbles, and for the Plateau-Rayleigh instability of liquid jets.

Journal Article↗
PubMed2003

DNA unzipped under a constant force exhibits multiple metastable intermediates.

Single molecule studies, at constant force, of the separation of double-stranded DNA into two separated single strands may provide information relevant to the dynamics of DNA replication. At constant applied force, theory predicts that the unzipped length as a function of time is characterized by jumps during which the strands separate rapidly, followed by long pauses where the number of separated base pairs remains constant. Here, we report previously uncharacterized observations of this striking behavior carried out on a number of identical single molecules simultaneously. When several single lambda phage molecules are subject to the same applied force, the pause positions are reproducible in each. This reproducibility shows that the positions and durations of the pauses in unzipping provide a sequence-dependent molecular fingerprint. For small forces, the DNA remains in a partially unzipped state for at least several hours. For larger forces, the separation is still characterized by jumps and pauses, but the double-stranded DNA will completely unzip in less than 30 min.

Adsorption↗
PubMed2003

Comparison of P450s from human and fugu: 420 million years of vertebrate P450 evolution.

The fugu (pufferfish) genome has been sequenced, and a second genome assembly was released 17 May 2002. Exhaustive searches were made to identify all P450 genes and pseudogenes from the earlier release of 26 October 2001. P450 genes assembled as completely as possible from these data were used to do additional searches of the newer assembly and all P450 genes and pseudogenes in the available fugu sequence data have been identified, compared to human P450s, and assigned names. There are 54 P450 genes in fugu and 1 nearly intact pseudogene (CYP3A50P). CYP1A is missing much of its N-terminal half; however, 45 P450 genes are completely assembled. Eight others are lacking only one or two exons or less. CYP2X4 is known only from an EST. This may be a 55th P450 gene if it represents an accurate sequence. In addition to 2X4, there are 16 other pseudogene fragments or small pieces of P450 genes. At the P450 family level, 17 of 18 mammalian families are found in fugu. CYP39 is the only CYP family missing and it is not seen in any other fish sequence data either. The CYP2 family shows the largest degree of divergence. In the CYP2 family, only CYP2R1 and CYP2U1 are conserved as recognizable subfamilies across species. Intron-exon boundaries are largely preserved across 420 million years of evolution.

Algorithms↗
PubMed2003

Sexual dysfunction in men undergoing infertility evaluation: a cohort observational study.

OBJECTIVE: To study psychosexual problems in men undergoing infertility evaluation. DESIGN: A cohort observational study. SETTING: Male infertility diagnostic center. PATIENT(S): Four hundred twelve men undergoing infertility evaluation between 1999 and 2001. INTERVENTION(S): Baseline and follow-up data on sexual functions. Semen analysis for samples collected by masturbation. A second analysis was requested in 2 weeks upon finding an abnormality of semen parameters. Penile vibratory stimulation to help men who failed to collect semen on their second or subsequent appointments. MAIN OUTCOME MEASURE(S): Sexual functions (erection and orgasm) during the time of infertility evaluation. RESULT(S): Seven of 412 men were excluded from the analysis due to a past history of sexual dysfunction. Of the remaining 405 men, 46 (11%) failed to collect semen by masturbation for a second semen analysis after repeated (2-4 times) attempts at 2- to 3-day intervals. Nine of these men (20%) were able to collect semen using vibratory stimulation. All 46 men experienced problems with erection or orgasm and had severe anxiety during attempts to masturbate and during sexual contact with their partners. CONCLUSION(S): Our study indicates that some men may experience sexual dysfunction of a psychogenic nature in response to the diagnosis of infertility.

Adult↗
PubMed2003

Negative effects of increased sperm DNA damage in relation to seminal oxidative stress in men with idiopathic and male factor infertility.

OBJECTIVE: To examine the effects of increased sperm DNA damage in relation to seminal oxidative stress in men with idiopathic and male factor infertility. Prospective study. SETTINGS: Infertility clinic at a tertiary care academic institution. PATIENT(S): Ninety-two infertile men with normal female partners. Sixteen fertile donors served as the control group. INTERVENTION(S): Standard semen analysis and assessment of levels of seminal oxidative stress. Assisted reproductive techniques in 33 of the 92 patients (IUI [n = 19], IVF [n = 10], and intracytoplasmic sperm injection [n = 4]). MAIN OUTCOME MEASURE(S): Sperm DNA damage by sperm chromatin structure assay. Results were expressed as DNA fragmentation index. RESULT(S): Patients were classified as having either idiopathic (n = 23) or male factor infertility (n = 69). Patients with idiopathic and male factor infertility had significantly higher DNA fragmentation index and oxidative stress compared with the case of fertile donors. A clinical pregnancy was achieved in 9 (27%) of 33 patients with assisted reproductive techniques. Significantly higher DNA fragmentation index and oxidative stress were found in men who failed to initiate a pregnancy after assisted reproductive techniques (n = 24), compared with the cases of those who succeeded and of the fertile donors. DNA fragmentation index was correlated positively with oxidative stress (r = 0.27), and negatively with fertilization (r = -0.70) and embryo quality (r = -0.70). CONCLUSION(S): Sperm DNA damage is significantly increased in men with idiopathic and male factor infertility and in men who failed to initiate a pregnancy after assisted reproductive techniques. Such an increase may be related to high levels of seminal oxidative stress.

Adult↗
PubMed2003

Acute posttraumatic planovalgus foot deformity involving hindfoot ligamentous pathology.

This article outlines the pathology of posttraumatic adult flat foot deformity that involves hindfoot ligamentous injury. The clinical presentation, radiographic findings, and pathology that are noted at the time of reconstruction are described. A proposed surgical reconstruction is presented that has successfully reconstructed the longitudinal arch of the affected foot.

Acute Disease↗
PubMed2003

Gene expression associated with interferon alfa antiviral activity in an HCV replicon cell line.

Interferon alfa (IFN-alpha)-based treatment is the only therapeutic option for chronic hepatitis C viral infection. However, the molecular mechanisms of IFN-alpha antiviral activity are not completely understood. The recent development of an HCV replicon cell culture system provides a feasible experimental model to investigate the molecular details of IFN-induced direct antiviral activity in hepatocytes. In this report, we show that IFN-alpha can effectively inhibit HCV subgenomic RNA replication and suppress viral nonstructural protein synthesis. Using cDNA microarray analysis, we also show that the replicon cells have different gene expression profile compared with the parental hepatoma cells (Huh7). IFN-alpha can induce a number of responsive genes in the replicon cells. One of the genes, 6-16 (G1P3), can enhance IFN-alpha antiviral efficacy. In addition, we demonstrate that IFN-alpha can significantly activate STAT3 in hepatoma cells, suggesting that this pathway plays a role in IFN-alpha signaling. In conclusion, our results indicate that IFN-alpha antiviral activity is associated with activation of STAT3-signaling pathway and intracellular gene activation. Our results also suggest that IFN-alpha-induced target genes may play an important role in IFN-alpha anti-HCV activity.

Antiviral Agents↗
PubMed2003

CD8+ T-cell interaction with HCV replicon cells: evidence for both cytokine- and cell-mediated antiviral activity.

The interaction between the host immune response and infected hepatocytes plays a central role in the pathogenesis of hepatitis C virus (HCV). The lack of a suitable animal or in vitro model has hindered our understanding of the host T-cell/HCV interaction. Our aim was to develop an in vitro model to study the mechanisms of HCV-specific T-cell-mediated antiviral and cytolytic function. The HCV replicon was HLA typed and lymphocytes were obtained from an HLA class I-matched subject. CD8(+) T cells were expanded with 2 HCV-specific/HLA-restricted peptides for NS3. Lymphocyte preparations were cocultured with HCV replicon (FCA1) and control (Huh7) cells labeled with (51)Cr. After a 48-hour incubation, the cells were harvested for RNA extraction. Standard blocking assays were performed in the presence of anti-interferon gamma (IFN-gamma), anti-tumor necrosis factor alpha (TNF-alpha), and anti-FasL. Cytolytic activity was measured by (51)Cr release. HCV replicon cells express homozygous HLA-A11 alleles and present HCV nonstructural proteins. HCV-specific expansion of CD8(+) cells led to a 10-fold decrease in HCV replication by Northern blot analysis and 21% specific lysis of FCA1 cells (compared with 2% of control Huh7 cells). Twenty percent of this antiviral activity was independent of T-cell binding, suggesting cytokine-mediated antiviral activity. The CD8(+) antiviral effect was markedly reduced by blocking either IFN-gamma or FasL but was unaffected by blocking TNF-alpha. In conclusion, HCV-specific CD8(+) cells inhibit viral RNA replication by cytokine-mediated and direct cytolytic effects. This T-cell/HCV subgenomic replicon system represents a model for the investigation of CD8 cell interaction with HCV-infected hepatocytes.

Antibodies↗