Correction: The extracellular Leucine-Rich Repeat superfamily; a comparative survey and analysis of evolutionary relationships and expression patterns

Correction to Dolan J, Walshe K, Alsbury S, Hokamp K, O'Keeffe S, Okafuji T, Miller SF, Tear G, Mitchell KJ: The extracellular leucine-rich repeat superfamily; a comparative survey and analysis of evolutionary relationships and expression patterns. BMC Genomics 2007, 8:320

Expression of endogenous Mkp1 in 6-OHDA rat models of Parkinson's disease.

We have previously demonstrated that mitogen-activated protein kinase phosphatase 1, Mkp1, is expressed in the developing and rat adult substantia nigra and striatum, where it promotes the growth of nigral dopaminergic neurons. Mkp1 may therefore have therapeutic potential for Parkinson's disease. In the present study, we have assessed the expression of Mkp1 and TH in the substantia nigra and striatum of parkinsonian rat models. Expression was measured at 4 and 10 days post-lesion in the 6-hydroxydopamine (6-OHDA) medial forebrain bundle lesion model and after 4, 10 and 28 days in the 6-OHDA striatal lesion model. Our results show that Mkp1 expression was transiently up-regulated in the substantia nigra at 4 days post-6-OHDA administration in the two models while TH expression was decreased at the later time-points examined. These data suggest that Mkp1 may play a role in counteracting the neurotoxic effects of 6-OHDA in nigral dopaminergic neurons

Zeb2 is a negative regulator of midbrain dopaminergic axon growth and target innervation

Neural connectivity requires neuronal differentiation, axon growth, and precise target innervation. Midbrain dopaminergic neurons project via the nigrostriatal pathway to the striatum to regulate voluntary movement. While the specification and differentiation of these neurons have been extensively studied, the molecular mechanisms that regulate midbrain dopaminergic axon growth and target innervation are less clear. Here we show that the transcription factor Zeb2 cell-autonomously represses Smad signalling to limit midbrain dopaminergic axon growth and target innervation. Zeb2 levels are downregulated in the embryonic rodent midbrain during the period of dopaminergic axon growth, when BMP pathway components are upregulated. Experimental knockdown of Zeb2 leads to an increase in BMP-Smad-dependent axon growth. Consequently there is dopaminergic hyperinnervation of the striatum, without an increase in the numbers of midbrain dopaminergic neurons, in conditional Zeb2 (Nestin-Cre based) knockout mice. Therefore, these findings reveal a new mechanism for the regulation of midbrain dopaminergic axon growth during central nervous system development

Lymph node homing cells biologically enriched for γδ T cells express multiple genes from the T19 repertoire

Sheep γδ T cells have been shown serologically to express T19, a membrane protein of 180-200 kDa which is a member of the scavenger receptor superfamily. Previous work from this laboratory resulted in the detection of a multigene family of T19-like genes in the sheep genome. In this study nucleotide sequences from several T19 genes were determined and are reported along with the corresponding segments of a number of expressed mRNA molecules. A segment of a single sheep T19-like gene was sequenced and these data, along with the corresponding sequences from cloned T19-like cDNA molecules from sheep and cow, were used to design an ollgonucleotide primer system suitable for amplification of corresponding segments of many T19 genes and their cDNAs. Between 30 and 40% of cloned T19 genes were amenable to amplification using the selected primers, and sequence analysis of cloned PCR products confirmed that different T19 genes encode unique amino acid sequences. The expression of multiple T19 genes was established using cDNA molecules obtained from a single sample of sheep lymphocyte mRNA. The possible role of the T19 family of genes is discusse

Exposure of the yeast Candida albicans to the anti-neoplastic agent adriamycin increases the tolerance to amphotericin B

Cancer patients experience a high incidence of fungal infections due to their immuno-suppressed condition. This work has investigated the interaction of an anti-neoplastic agent, adriamycin (doxorubicin), with the yeast Candida albicans and examined whether this drug altered the susceptibility of the yeast to amphotericin B - an anti- fungal agent used for the treatment of systemic fungal infections in cancer patients. Exposure to adriamycin for 24 h increased the growth of C. albicans and increased the tolerance to amphotericin B by a small, but statistically significant, extent. Growth in adriamycin-supplemented medium suppressed the respiration rate of C. albicans, which resulted in a decrease in the ergosterol content of the fungal cell membrane. The tolerance to amphotericin B was lost after exposure to adriamycin for 48 h, which coincided with a restoration in the respiration rate and the ergosterol content of the fungal cell membrane. This work demonstrated that short-term exposure (24 h) to adriamycin increased the tolerance of C. albicans for amphotericin B, which may be mediated by a decrease in the ergosterol content as a result of an adriamycin-induced disruption of oxidative phosphorylation

TNF superfamily member APRIL enhances midbrain dopaminergic axon growth and contributes to the nigrostriatal projection in vivo

We have studied the role of the tumor necrosis factor superfamily member APRIL in the development of embryonic mouse midbrain dopaminergic neurons in vitro and in vivo. In culture, soluble APRIL enhanced axon growth during a window of development between E12 and E14 when nigrostriatal axons are growing to their targets in the striatum in vivo. April transcripts were detected in both the striatum and midbrain during this period and at later stages. The axon growth–enhancing effect of APRIL was similar to that of glial cell-derived neurotrophic factor (GDNF), but in contrast to GDNF, APRIL did not promote the survival of midbrain dopaminergic neurons. The effect of APRIL on axon growth was prevented by function-blocking antibodies to one of its receptors, BCMA (TNFRSF13A), but not by function-blocking antibodies to the other APRIL receptor, TACI (TNFRSF13B), suggesting that the effects of APRIL on axon growth are mediated by BCMA. In vivo, there was a significant reduction in the density of midbrain dopaminergic projections to the striatum in April −/− embryos compared with wild type littermates at E14. These findings demonstrate that APRIL is a physiologically relevant factor for the nigrostriatal projection. Given the importance of the degeneration of dopaminergic nigrostriatal connections in the pathogenesis and progression of Parkinson's disease, our findings contribute to our understanding of the factors that establish nigrostriatal integrity.</p

The Long-Baseline Neutrino Experiment: Exploring Fundamental Symmetries of the Universe

The preponderance of matter over antimatter in the early Universe, the dynamics of the supernova bursts that produced the heavy elements necessary for life and whether protons eventually decay --- these mysteries at the forefront of particle physics and astrophysics are key to understanding the early evolution of our Universe, its current state and its eventual fate. The Long-Baseline Neutrino Experiment (LBNE) represents an extensively developed plan for a world-class experiment dedicated to addressing these questions. LBNE is conceived around three central components: (1) a new, high-intensity neutrino source generated from a megawatt-class proton accelerator at Fermi National Accelerator Laboratory, (2) a near neutrino detector just downstream of the source, and (3) a massive liquid argon time-projection chamber deployed as a far detector deep underground at the Sanford Underground Research Facility. This facility, located at the site of the former Homestake Mine in Lead, South Dakota, is approximately 1,300 km from the neutrino source at Fermilab -- a distance (baseline) that delivers optimal sensitivity to neutrino charge-parity symmetry violation and mass ordering effects. This ambitious yet cost-effective design incorporates scalability and flexibility and can accommodate a variety of upgrades and contributions. With its exceptional combination of experimental configuration, technical capabilities, and potential for transformative discoveries, LBNE promises to be a vital facility for the field of particle physics worldwide, providing physicists from around the globe with opportunities to collaborate in a twenty to thirty year program of exciting science. In this document we provide a comprehensive overview of LBNE's scientific objectives, its place in the landscape of neutrino physics worldwide, the technologies it will incorporate and the capabilities it will possess.Comment: Major update of previous version. This is the reference document for LBNE science program and current status. Chapters 1, 3, and 9 provide a comprehensive overview of LBNE's scientific objectives, its place in the landscape of neutrino physics worldwide, the technologies it will incorporate and the capabilities it will possess. 288 pages, 116 figure

The extracellular Leucine-rich repeat superfamily; a comparative survey and analysis of evolutionary relationships and expression patterns

Background: Leucine-rich repeats (LRRs) are highly versatile and evolvable protein-ligand interaction motifs found in a large number of proteins with diverse functions, including innate immunity and nervous system development. Here we catalogue all of the extracellular LRR (eLRR) proteins in worms, flies, mice and humans. We use convergent evidence from several transmembrane-prediction and motif-detection programs, including a customised algorithm, LRRscan, to identify eLRR proteins, and a hierarchical clustering method based on TribeMCL to establish their evolutionary relationships. Results: This yields a total of 369 proteins (29 in worm, 66 in fly, 135 in mouse and 139 in human), many of them of unknown function. We group eLRR proteins into several classes: those with only LRRs, those that cluster with Toll-like receptors (Tlrs), those with immunoglobulin or fibronectin-type 3 (FN3) domains and those with some other domain. These groups show differential patterns of expansion and diversification across species. Our analyses reveal several clusters of novel genes, including two Elfn genes, encoding transmembrane proteins with eLRRs and an FN3 domain, and six genes encoding transmembrane proteins with eLRRs only (the Elron cluster). Many of these are expressed in discrete patterns in the developing mouse brain, notably in the thalamus and cortex. We have also identified a number of novel fly eLRR proteins with discrete expression in the embryonic nervous system. Conclusion: This study provides the necessary foundation for a systematic analysis of the functions of this class of genes, which are likely to include prominently innate immunity, inflammation and neural development, especially the specification of neuronal connectivity

A proteomic approach to investigating gene cluster expression and secondary metabolite functionality in Aspergillus fumigatus.

A combined proteomics and metabolomics approach was utilised to advance the identification and characterisation of secondary metabolites in Aspergillus fumigatus. Here, implementation of a shotgun proteomic strategy led to the identification of non-redundant mycelial proteins (n = 414) from A. fumigatus including proteins typically under-represented in 2-D proteome maps: proteins with multiple transmembrane regions, hydrophobic proteins and proteins with extremes of molecular mass and pI. Indirect identification of secondary metabolite cluster expression was also achieved, with proteins (n = 18) from LaeA-regulated clusters detected, including GliT encoded within the gliotoxin biosynthetic cluster. Biochemical analysis then revealed that gliotoxin significantly attenuates H2O2-induced oxidative stress in A. fumigatus (p>0.0001), confirming observations from proteomics data. A complementary 2-D/LC-MS/MS approach further elucidated significantly increased abundance (p<0.05) of proliferating cell nuclear antigen (PCNA), NADH-quinone oxidoreductase and the gliotoxin oxidoreductase GliT, along with significantly attenuated abundance (p<0.05) of a heat shock protein, an oxidative stress protein and an autolysis-associated chitinase, when gliotoxin and H2O2 were present, compared to H2O2 alone. Moreover, gliotoxin exposure significantly reduced the abundance of selected proteins (p<0.05) involved in de novo purine biosynthesis. Significantly elevated abundance (p<0.05) of a key enzyme, xanthine-guanine phosphoribosyl transferase Xpt1, utilised in purine salvage, was observed in the presence of H2O2 and gliotoxin. This work provides new insights into the A. fumigatus proteome and experimental strategies, plus mechanistic data pertaining to gliotoxin functionality in the organism

Development of an in vitro media perfusion model of Leishmania major macrophage infection.

BACKGROUND: In vitro assays are widely used in studies on pathogen infectivity, immune responses, drug and vaccine discovery. However, most in vitro assays display significant differences to the in vivo situation and limited predictive properties. We applied medium perfusion methods to mimic interstitial fluid flow to establish a novel infection model of Leishmania parasites. METHODS: Leishmania major infection of mouse peritoneal macrophages was studied within the Quasi Vivo QV900 macro-perfusion system. Under a constant flow of culture media at a rate of 360μl/min, L. major infected macrophages were cultured either at the base of a perfusion chamber or raised on 9mm high inserts. Mathematical and computational modelling was conducted to estimate medium flow speed, shear stress and oxygen concentration. The effects of medium flow on infection rate, intracellular amastigote division, macrophage phagocytosis and macropinocytosis were measured. RESULTS: Mean fluid speeds at the macrophage cell surface were estimated to be 1.45 x 10-9 m/s and 1.23 x 10-7 m/s for cells at the base of the chamber and cells on an insert, respectively. L. major macrophage infection was significantly reduced under both media perfusion conditions compared to cells maintained under static conditions; a 85±3% infection rate of macrophages at 72 hours in static cultures compared to 62±5% for cultures under slow medium flow and 55±3% under fast medium flow. Media perfusion also decreased amastigote replication and both macrophage phagocytosis (by 44±4% under slow flow and 57±5% under fast flow compared with the static condition) and macropinocytosis (by 40±4% under slow flow and 62±5% under fast flow compared with the static condition) as measured by uptake of latex beads and pHrodo Red dextran. CONCLUSIONS: Perfusion of culture medium in an in vitro L. major macrophage infection model (simulating in vivo lymphatic flow) reduced the infection rate of macrophages, the replication of the intracellular parasite, macrophage phagocytosis and macropinocytosis with greater reductions achieved under faster flow speeds