Biomedical subjects

David Horn

Publications and source records attributed to David Horn.

At least 19 recordsLinked to original sources

PubMed2026

Zinc-dependent turnover of ZIP3 transporter mRNA by trypanosome ZNK1.

Like other cells, parasitic and other trypanosomatids sense Zn2+ and regulate Zn2+ transport, but the mechanisms involved remained unknown. Here, we identify a trypanosome RNA-binding protein that specifically eliminates ZIP3 transporter mRNA in Zn2+-replete conditions. We first demonstrate that Trypanosoma brucei ZIP3 mRNA abundance is subject to 3'-untranslated region (3'-UTR) and Zn2+-dependent negative control. A genome-wide RNA interference library screen, using a reporter associated with the ZIP3 3'-UTR, identifies Tb927.11.9510 as a candidate Zn2+-sensor, and we name this protein Zinc Nuclear Knuckles 1 (ZNK1) since it localizes to the nucleus and contains several Zn2+-knuckle motifs. ZNK1 is conserved among trypanosomatids, and a PIN domain suggests a ribonuclease-based mechanism. We use Cas9-editing to knockout ZNK1 and observe specific accumulation of ZIP3 transcripts, and increased intracellular Zn2+, in znk1 null cells. We validate ZNK1 as a ZIP3 3'-UTR-dependent negative regulator and identify a GU-repeat motif in the ZIP3 3'-UTR that is predictive of ZNK1-based negative control. In conclusion, ZNK1 eliminates ZIP3 transporter mRNA in a Zn2+-dependent manner. We suggest that trypanosomatid ZNK1 is a highly selective zinc finger nuclease that binds GU-repeat motifs within ZIP3 3'-UTRs and degrades Zn2+ transporter mRNA only when the tandem sensor knuckle modules are coordinated with Zn2+.

Trypanosoma brucei brucei↗

PubMed2007

A sirtuin in the African trypanosome is involved in both DNA repair and telomeric gene silencing but is not required for antigenic variation.

Silent information regulator 2 (Sir2)-related proteins or sirtuins function as NAD(+)-dependent deacetylases or ADP ribosylases that target a range of substrates, thereby influencing chromatin structure and a diverse range of other biological functions. Genes encoding three Sir2-related proteins (SIR2rp1-3) have been identified in the parasitic trypanosomatids, early branching protozoa with no previously reported transcriptional silencing machinery. Here we show that, in the mammalian-infective bloodstream-stage of the African trypanosome, Trypanosoma brucei, SIR2rp1 localizes to the nucleus while SIR2rp2 and SIR2rp3 are both mitochondrial proteins. The nuclear protein, SIR2rp1, controls DNA repair and repression of RNA polymerase I-mediated expression immediately adjacent to telomeres. Antigenic variation, however, which involves the silencing and Pol I-mediated transcriptional switching of subtelomeric variant surface glycoprotein genes, continues to operate independent of SIR2rp1.

Amino Acid Sequence↗

PubMed2006

Deletion of the Trypanosoma brucei superoxide dismutase gene sodb1 increases sensitivity to nifurtimox and benznidazole.

It has been more than 25 years since it was first reported that nifurtimox and benznidazole promote superoxide production in trypanosomes. However, there has been no direct evidence of an association between the drug-induced free radicals and trypanocidal activity. Here, we identify a superoxide dismutase required to protect Trypanosoma brucei from drug-generated superoxide.

Animals↗

PubMed2006

Novel unsupervised feature filtering of biological data.

MOTIVATION: Many methods have been developed for selecting small informative feature subsets in large noisy data. However, unsupervised methods are scarce. Examples are using the variance of data collected for each feature, or the projection of the feature on the first principal component. We propose a novel unsupervised criterion, based on SVD-entropy, selecting a feature according to its contribution to the entropy (CE) calculated on a leave-one-out basis. This can be implemented in four ways: simple ranking according to CE values (SR); forward selection by accumulating features according to which set produces highest entropy (FS1); forward selection by accumulating features through the choice of the best CE out of the remaining ones (FS2); backward elimination (BE) of features with the lowest CE. RESULTS: We apply our methods to different benchmarks. In each case we evaluate the success of clustering the data in the selected feature spaces, by measuring Jaccard scores with respect to known classifications. We demonstrate that feature filtering according to CE outperforms the variance method and gene-shaving. There are cases where the analysis, based on a small set of selected features, outperforms the best score reported when all information was used. Our method calls for an optimal size of the relevant feature set. This turns out to be just a few percents of the number of genes in the two Leukemia datasets that we have analyzed. Moreover, the most favored selected genes turn out to have significant GO enrichment in relevant cellular processes.

Artificial Intelligence↗

PubMed2006

Bloodstream infections in hospitalized adults with sickle cell disease: a retrospective analysis.

Bloodstream infections (BSI) are a common cause of morbidity and mortality in people with sickle cell disease (SCD). In children with SCD, BSI are most often caused by encapsulated organisms. There is a surprising paucity of medical literature that is focused on evaluating SCD adults with BSI. We reviewed the charts of adults with SCD and BSI who were admitted to our hospital between April 1999 and August 2003. During this period a total of 1,692 hospital admissions for 193 adults with SCD were identified and 28% of these patients had at least 1 episode of positive blood cultures, with 69 episodes (17%) considered true BSI. Nosocomial BSI occurred in 34 episodes (49%). Among community BSI, in contrast to BSI in children with SCD, Streptococcus pneumoniae was rarely encountered. A high incidence of staphylococcal BSI in adults with SCD was noted. Twenty-eight percent of all BSI were caused by Staphylococcus aureus, and 15 of 22 isolates (68%) of these were methicillin-resistant. Gram-negative organisms, anaerobes, and yeast were found in 21 (27%), 3 (4%), and 4 isolates (5%) of BSI, respectively. Since over 80% of BSI were considered catheter-related, the higher incidence of gram-positive bacterial infections was likely due to the presence of indwelling central venous catheters. Empiric therapy for adults with SCD suspected of having BSI, especially in the presence of indwelling central venous catheters, should include antimicrobial therapy targeted at gram-positive bacteria (especially MRSA) and gram-negative bacteria. Also, if patients are critically ill, consideration should be made to include antifungal agents. Additional research into the adult SCD population appears necessary to further define this problem.

Adult↗

PubMed2006

Proteomics and Beyond: a report on the 3rd Annual Spring Workshop of the HUPO-PSI 21-23 April 2006, San Francisco, CA, USA.

The theme of the third annual Spring workshop of the HUPO-PSI was "proteomics and beyond" and its underlying goal was to reach beyond the boundaries of the proteomics community to interact with groups working on the similar issues of developing interchange standards and minimal reporting requirements. Significant developments in many of the HUPO-PSI XML interchange formats, minimal reporting requirements and accompanying controlled vocabularies were reported, with many of these now feeding into the broader efforts of the Functional Genomics Experiment (FuGE) data model and Functional Genomics Ontology (FuGO) ontologies.

Animals↗

PubMed2006

A case of recurrent episodes of Candida parapsilosis fungemia.

Candida species is the fourth most commonly isolated organism in blood stream infections in the hospital setting. Patients with candidemia frequently succumb to this infection. For those that survive an initial candidemia, an increasing number of cases of breakthrough or recurrent candidemias have been reported in the literature. We report three episodes of C. parapsilosis fungemia in a cancer patient. Molecular testing was performed and confirmed that all these episodes occurring within an eight-month period were caused by the same organism. The incidence of recurrent candidemia is likely to increase in the near future. Studies and therapeutic interventions for patients at risk for recurrent candidemias are warranted.

Adenocarcinoma↗

PubMed2006

Repression of polymerase I-mediated gene expression at Trypanosoma brucei telomeres.

The African trypanosome, Trypanosoma brucei, is a flagellated pathogenic protozoan that branched early from the eukaryotic lineage. Unusually, it uses RNA polymerase I (Pol I) for mono-telomeric expression of variant surface glycoprotein (VSG) genes in bloodstream-form cells. Many other subtelomeric VSG genes are reversibly repressed, but no repressive DNA sequence has been identified in any trypanosomatid. Here, we show that artificially seeded de novo telomeres repress Pol I-dependent gene expression in mammalian bloodstream and insect life-cycle stages of T. brucei. In a telomeric VSG expression site, repression spreads further along the chromosome and this effect is specific to the bloodstream stage. We also show that de novo telomere extension is telomerase dependent and that the rate of extension correlates with the expression level of the adjacent gene. Our results show constitutive telomeric repression in T. brucei and indicate that an enhanced, developmental stage-specific repression mechanism controls antigenic variation.

Animals↗

PubMed2006

Chromosome-wide analysis of gene function by RNA interference in the african trypanosome.

Trypanosomatids of the order Kinetoplastida are major contributors to global disease and morbidity, and understanding their basic biology coupled with the development of new drug targets represents a critical need. Additionally, trypanosomes are among the more accessible divergent eukaryote experimental systems. The genome of Trypanosoma brucei contains 8,131 predicted open reading frames (ORFs), of which over half have no known homologues beyond the Kinetoplastida and a substantial number of others are poorly defined by in silico analysis. Thus, a major challenge following completion of the T. brucei genome sequence is to obtain functional data for all trypanosome ORFs. As T. brucei is more experimentally tractable than the related Trypanosoma cruzi and Leishmania spp. and shares >75% of their genes, functional analysis of T. brucei has the potential to inform a range of parasite biology. Here, we report methods for systematic mRNA ablation by RNA interference (RNAi) and for phenotypic analysis, together with online data dissemination. This represents the first systematic analysis of gene function in a parasitic organism. In total, 210 genes have been targeted in the bloodstream form parasite, representing an essentially complete phenotypic catalogue of chromosome I together with a validation set. Over 30% of the chromosome I genes generated a phenotype when targeted by RNAi; most commonly, this affected cell growth, viability, and/or cell cycle progression. RNAi against approximately 12% of ORFs was lethal, and an additional 11% had growth defects but retained short-term viability in culture. Although we found no evidence for clustering or a bias towards widely evolutionarily conserved genes within the essential ORF cohort, the putative chromosome I centromere is adjacent to a domain containing genes with no associated phenotype. Involvement of such a large proportion of genes in robust growth in vitro indicates that a high proportion of the expressed trypanosome genome is required for efficient propagation; many of these gene products represent potential drug targets.

Animals↗

PubMed2006

Complications of central venous catheters used for the treatment of acute hematogenous osteomyelitis.

OBJECTIVE: To determine the complications and risk factors for complications associated with using central venous catheters (CVCs) for the treatment of acute hematogenous osteomyelitis (AHO). METHODS: We conducted a retrospective cohort study of all patients admitted to the Children's Hospital of Philadelphia between January 1, 2000, and December 31, 2003, with a diagnosis of AHO. RESULTS: Eighty patients with AHO met inclusion criteria. The median age was 5 years, and 66% of the patients were male. The most commonly affected bones were the femur (25%), tibia (20%), and pelvis (16%). Staphylococcus aureus was the most common organism identified from cultures of bone (67%) and blood (30%). Seventy-five patients (94%) received >2 weeks of intravenous (IV) antibiotic therapy via a CVC and 5 (6%) received <2 weeks of IV antibiotic therapy before conversion to oral therapy for a median of 25 days. None of the patients who switched to oral therapy within 2 weeks was rehospitalized or returned to the emergency department. Of the 75 patients who received >2 weeks of IV therapy, 41% had > or =1 CVC-associated complication. Seventeen patients (23%) had a CVC malfunction or displacement, 8 (11%) had a catheter-associated bloodstream infection, 8 (11%) had fever with negative blood culture results, and 4 (5%) had a local skin infection at the site of catheter insertion. Older age was protective against the development of a CVC-associated complication, whereas the lowest median household income was associated with development of a CVC-associated complication. CONCLUSIONS: Interventions to reduce CVC-associated complications should be developed and evaluated, particularly for young children and those from families with low household incomes. Clinical trials are needed to evaluate the safety and efficacy of oral antibiotic therapy after a short course of IV therapy as an alternative to prolonged IV therapy.

Acute Disease↗

PubMed2005

Tagging a T. brucei RRNA locus improves stable transfection efficiency and circumvents inducible expression position effects.

In Trypanosoma brucei, RNA interference (RNAi) and recombinant protein expression are established as powerful approaches for functional genomics, particularly when combined with inducible expression. The favoured methods involve exploiting homologous recombination to target expression cassettes to a chromosome sub-set to establish stable cell lines. Unfortunately, bloodstream-form cells, those that cause disease in mammals, exhibit low efficiency stable transfection. Current expression systems can also exhibit other undesirable features, including variable position effects and leaky, inducible expression. We have developed systems in bloodstream-form cells that alleviate these problems. Using constructs for RNAi and expression of (GFP) tagged proteins, we target a (hyg) tagged ribosomal RNA (RRNA) locus which circumvents position effects and allows increased targeting efficiency. We also report a compatible double-inducible system for tight regulation of highly toxic products. This system exploits a new inducible RRNA promoter to drive T7 RNA polymerase (T7RNAP) transcription which then drives expression from inducible T7 promoters. The developments described should facilitate functional analysis and increased throughput.

Animals↗

PubMed2005

Functional characterisation of the iron superoxide dismutase gene repertoire in Trypanosoma brucei.

Superoxide dismutases (SOD) are a family of antioxidant enzymes that function by removing superoxide anions from the cellular environment. Here, we show that the African trypanosome, Trypanosoma brucei, expresses four SOD isoforms, three of which we have validated biochemically as iron dependent, a feature normally associated with prokaryotic SODs. Localisation studies reveal that two of the enzymes are found predominantly in a parasite-specific organelle, the glycosome (TbSODB1 and TbSODB2), while the other two are targeted to the mitochondrion (TbSODA and TbSODC). Functional analysis of the SOD repertoire in bloodstream form parasites was performed using an inducible RNA interference (RNAi) approach. Down-regulation of the glycosomal SOD transcripts corresponded with a significant reduction in the corresponding proteins and a dramatic level of cell death within the population. The importance of one of the mitochondrial enzymes (TbSODA) only became apparent when parasites were exposed to the superoxide-generating agent paraquat following induction of RNAi. These experiments therefore identify essential components of the superoxide metabolising arm of the T. brucei oxidative defence system and validate these enzymes as parasite-specific targets for drug design.

Amino Acid Sequence↗

PubMed2005

Unsupervised learning of natural languages.

We address the problem, fundamental to linguistics, bioinformatics, and certain other disciplines, of using corpora of raw symbolic sequential data to infer underlying rules that govern their production. Given a corpus of strings (such as text, transcribed speech, chromosome or protein sequence data, sheet music, etc.), our unsupervised algorithm recursively distills from it hierarchically structured patterns. The adios (automatic distillation of structure) algorithm relies on a statistical method for pattern extraction and on structured generalization, two processes that have been implicated in language acquisition. It has been evaluated on artificial context-free grammars with thousands of rules, on natural languages as diverse as English and Chinese, and on protein data correlating sequence with function. This unsupervised algorithm is capable of learning complex syntax, generating grammatical novel sentences, and proving useful in other fields that call for structure discovery from raw data, such as bioinformatics.

Algorithms↗

PubMed2005

Vitamin C biosynthesis in trypanosomes: a role for the glycosome.

The capacity to synthesize vitamin C (ascorbate) is widespread in eukaryotes but is absent from humans. The last step in the biosynthetic pathway involves the conversion of an aldonolactone substrate to ascorbate, a reaction catalyzed by members of an FAD-dependent family of oxidoreductases. Here we demonstrate that both the African trypanosome, Trypanosoma brucei, and the American trypanosome, Trypanosoma cruzi, have the capacity to synthesize vitamin C and show that this reaction occurs in a unique single-membrane organelle, the glycosome. The corresponding T. brucei flavoprotein (TbALO) obeys Michaelis-Menten kinetics and can utilize both L-galactono-gamma-lactone and D-arabinono-gamma-lactone as substrate, properties characteristic of plant and fungal enzymes. We could detect no activity toward the mammalian enzyme substrate L-gulono-gamma-lactone. TbALO null mutants (bloodstream form) were found to display a transient growth defect, a trait that was enhanced when they were cultured in medium in which the essential serum component had been pretreated with ascorbate oxidase to deplete vitamin C. It is implicit, therefore, that bloodstream-form trypanosomes also possess a capacity for ascorbate transport.

Amino Acid Sequence↗

PubMed2005

The genome sequence of Trypanosoma cruzi, etiologic agent of Chagas disease.

Whole-genome sequencing of the protozoan pathogen Trypanosoma cruzi revealed that the diploid genome contains a predicted 22,570 proteins encoded by genes, of which 12,570 represent allelic pairs. Over 50% of the genome consists of repeated sequences, such as retrotransposons and genes for large families of surface molecules, which include trans-sialidases, mucins, gp63s, and a large novel family (>1300 copies) of mucin-associated surface protein (MASP) genes. Analyses of the T. cruzi, T. brucei, and Leishmania major (Tritryp) genomes imply differences from other eukaryotes in DNA repair and initiation of replication and reflect their unusual mitochondrial DNA. Although the Tritryp lack several classes of signaling molecules, their kinomes contain a large and diverse set of protein kinases and phosphatases; their size and diversity imply previously unknown interactions and regulatory processes, which may be targets for intervention.

Animals↗

PubMed2005

The genome of the kinetoplastid parasite, Leishmania major.

Leishmania species cause a spectrum of human diseases in tropical and subtropical regions of the world. We have sequenced the 36 chromosomes of the 32.8-megabase haploid genome of Leishmania major (Friedlin strain) and predict 911 RNA genes, 39 pseudogenes, and 8272 protein-coding genes, of which 36% can be ascribed a putative function. These include genes involved in host-pathogen interactions, such as proteolytic enzymes, and extensive machinery for synthesis of complex surface glycoconjugates. The organization of protein-coding genes into long, strand-specific, polycistronic clusters and lack of general transcription factors in the L. major, Trypanosoma brucei, and Trypanosoma cruzi (Tritryp) genomes suggest that the mechanisms regulating RNA polymerase II-directed transcription are distinct from those operating in other eukaryotes, although the trypanosomatids appear capable of chromatin remodeling. Abundant RNA-binding proteins are encoded in the Tritryp genomes, consistent with active posttranscriptional regulation of gene expression.

Animals↗

PubMed2005

The influence of flotation agent concentration on the wettability and flotability of polystyrene.

The fundamental flotation process is the formation of a flocculant by air bubbles and solid particles in an aqueous solution. The behavior of plastic particles is significantly influenced by the wettability of the plastics. In this article the reciprocal relationship between the flotability and wettability of polystyrene was studied at different concentrations of flotation agents, particularly terpineol, polyethylene glycol dodecyl ether, tannic acid, and calcium lignosulfonate. The conclusions obtained demonstrate the dissimilar action of flotation depressants, what means different adhesion mechanisms on a plastic surface.

Journal Article↗

PubMed2005

The central roles of telomeres and subtelomeres in antigenic variation in African trypanosomes.

Telomeres and subtelomeres are important to the virulence of a number of pathogens, as they harbour large diverse gene families associated with the maintenance of infection. Evasion of immunity by African trypanosomes involves the differential expression of variant surface glycoproteins (VSGs), which are encoded by a family of >1500 genes and pseudogenes. This silent archive is located subtelomerically and is activated by gene conversion into specialized transcription units, which themselves are subject to silencing by allelic exclusion. Current research addresses the role of telomeres in the conversion and silencing mechanisms and in the diversification of the VSG archive.

Animals↗