A digital video system for observing and recording occultations

Stellar occultations by asteroids and outer solar system bodies can offer ground based observers with modest telescopes and camera equipment the opportunity to probe the shape, size, atmosphere and attendant moons or rings of these distant objects. The essential requirements of the camera and recording equipment are: good quantum efficiency and low noise, minimal dead time between images, good horological faithfulness of the image time stamps, robustness of the recording to unexpected failure, and low cost. We describe the Astronomical Digital Video occultation observing and recording System (ADVS) which attempts to fulfil these requirements and compare the system with other reported camera and recorder systems. Five systems have been built, deployed and tested over the past three years, and we report on three representative occultation observations: one being a 9 +/-1.5 second occultation of the trans-Neptunian object 28978 Ixion (mv=15.2) at 3 seconds per frame, one being a 1.51 +/-0.017 second occultation of Deimos, the 12~km diameter satellite of Mars, at 30 frames per second, and one being a 11.04 +/-0.4 second occultation, recorded at 7.5 frames per second, of the main belt asteroid, 361 Havnia, representing a low magnitude drop (Dmv = 0.4) occultation.Comment: 9 pages, 5 figures, 3 tables, accepted to Publications of the Astronomical Society of Australia (PASA

Truncation of TRIM5 in the <i>Feliformia</i> explains the absence of retroviral restriction in cells of the domestic cat

TRIM5[alpha] mediates a potent retroviral restriction phenotype in diverse mammalian species. Here, we identify a TRIM5 transcript in cat cells with a truncated B30.2 capsid binding domain and ablated restrictive function which, remarkably, is conserved across the &lt;i&gt;Feliformia&lt;/i&gt;. Cat TRIM5 displayed no restriction activity, but ectopic expression conferred a dominant negative effect against human TRIM5[alpha]. Our findings explain the absence of retroviral restriction in cat cells and suggest that disruption of the TRIM5 locus has arisen independently at least twice in the &lt;i&gt;Carnivora&lt;/i&gt;, with implications concerning the evolution of the host and pathogen in this taxon

Identification of cis-acting determinants mediating the unconventional secretion of tau.

The deposition of tau aggregates throughout the brain is a pathological characteristic within a group of neurodegenerative diseases collectively termed tauopathies, which includes Alzheimer's disease. While recent findings suggest the involvement of unconventional secretory pathways driving tau into the extracellular space and mediating the propagation of the disease-associated pathology, many of the mechanistic details governing this process remain elusive. In the current study, we provide an in-depth characterization of the unconventional secretory pathway of tau and identify novel molecular determinants that are required for this process. Here, using Drosophila models of tauopathy, we correlate the hyperphosphorylation and aggregation state of tau with the disease-related neurotoxicity. These newly established systems recapitulate all the previously identified hallmarks of tau secretion, including the contribution of tau hyperphosphorylation as well as the requirement for PI(4,5)P2 triggering the direct translocation of tau. Using a series of cellular assays, we demonstrate that both the sulfated proteoglycans on the cell surface and the correct orientation of the protein at the inner plasma membrane leaflet are critical determinants of this process. Finally, we identify two cysteine residues within the microtubule binding repeat domain as novel cis-elements that are important for both unconventional secretion and trans-cellular propagation of tau

Restriction of the felid lentiviruses by a synthetic feline TRIM5-CypA fusion

Gene therapy approaches to the treatment of HIV infection have targeted both viral gene expression and the cellular factors that are essential for virus replication. However, significant concerns have been raised regarding the potential toxic effects of such therapies, the emergence of resistant viral variants and unforeseen biological consequences such as enhanced susceptibility to unrelated pathogens. Novel restriction factors formed by the fusion of the tripartite motif protein (TRIM5) and cyclophilin A (CypA), or "TRIMCyps", offer an effective antiviral defence strategy with a very low potential for toxicity. In order to investigate the potential therapeutic utility of TRIMCyps in gene therapy for AIDS, a synthetic fusion protein between feline TRIM5 and feline CypA was generated and transduced into cells susceptible to infection with feline immunodeficiency virus (FIV). The synthetic feline TRIMCyp was highly efficient at preventing infection with both HIV and FIV and the cells resisted productive infection with FIV from either the domestic cat or the puma. Feline TRIMCyp and FIV infection of the cat offers a unique opportunity to evaluate TRIMCyp-based approaches to genetic therapy for HIV infection and the treatment of AIDS

Canine candidate genes for dilated cardiomyopathy: annotation of and polymorphic markers for 14 genes

BackgroundDilated cardiomyopathy is a myocardial disease occurring in humans and domestic animals and is characterized by dilatation of the left ventricle, reduced systolic function and increased sphericity of the left ventricle. Dilated cardiomyopathy has been observed in several, mostly large and giant, dog breeds, such as the Dobermann and the Great Dane. A number of genes have been identified, which are associated with dilated cardiomyopathy in the human, mouse and hamster. These genes mainly encode structural proteins of the cardiac myocyte.ResultsWe present the annotation of, and marker development for, 14 of these genes of the dog genome, i.e. alpha-cardiac actin, caveolin 1, cysteine-rich protein 3, desmin, lamin A/C, LIM-domain binding factor 3, myosin heavy polypeptide 7, phospholamban, sarcoglycan delta, titin cap, alpha-tropomyosin, troponin I, troponin T and vinculin. A total of 33 Single Nucleotide Polymorphisms were identified for these canine genes and 11 polymorphic microsatellite repeats were developed.ConclusionThe presented polymorphisms provide a tool to investigate the role of the corresponding genes in canine Dilated Cardiomyopathy by linkage analysis or association studies

A Method for the Acute and Rapid Degradation of Endogenous Proteins.

Methods for the targeted disruption of protein function have revolutionized science and greatly expedited the systematic characterization of genes. Two main approaches are currently used to disrupt protein function: DNA knockout and RNA interference, which act at the genome and mRNA level, respectively. A method that directly alters endogenous protein levels is currently not available. Here, we present Trim-Away, a technique to degrade endogenous proteins acutely in mammalian cells without prior modification of the genome or mRNA. Trim-Away harnesses the cellular protein degradation machinery to remove unmodified native proteins within minutes of application. This rapidity minimizes the risk that phenotypes are compensated and that secondary, non-specific defects accumulate over time. Because Trim-Away utilizes antibodies, it can be applied to a wide range of target proteins using off-the-shelf reagents. Trim-Away allows the study of protein function in diverse cell types, including non-dividing primary cells where genome- and RNA-targeting methods are limited

Copper(II)-Bis(Thiosemicarbazonato) Complexes as Antibacterial Agents: Insights into Their Mode of Action and Potential as Therapeutics

There is increasing interest in the use of lipophilic copper (Cu)-containing complexes to combat bacterial infections. In this work, we showed that Cu complexes with bis(thiosemicarbazone) ligands [Cu(btsc)] exert antibacterial activity against a range of medically significant pathogens. Previous work using Neisseria gonorrhoeae showed that Cu(btsc) complexes may act as inhibitors of respiratory dehydrogenases in the electron transport chain. We now show that these complexes are also toxic against pathogens that lack a respiratory chain. Respiration in Escherichia coli was slightly affected by Cu(btsc) complexes, but our results indicate that, in this model bacterium, the complexes act primarily as agents that deliver toxic Cu ions efficiently into the cytoplasm. Although the chemistry of Cu(btsc) complexes may dictate their mechanism of action, their efficacy depends heavily on bacterial physiology. This is linked to the ability of the target bacterium to tolerate Cu and, additionally, the susceptibility of the respiratory chain to direct inhibition by Cu(btsc) complexes. The physiology of N. gonorrhoeae, including multidrug-resistant strains, makes it highly susceptible to damage by Cu ions and Cu(btsc) complexes, highlighting the potential of Cu(btsc) complexes (and Cu-based therapeutics) as a promising treatment against this important bacterial pathogen

Microglia become hypofunctional and release metalloproteases and tau seeds when phagocytosing live neurons with P301S tau aggregates.

[Figure: see text]

Cytosolic Fc receptor TRIM21 inhibits seeded tau aggregation

Alzheimer's disease (AD) and other neurodegenerative disorders are associated with the cytoplasmic aggregation of microtubule-associated protein tau. Recent evidence supports transcellular transfer of tau misfolding (seeding) as the mechanism of spread within an affected brain, a process reminiscent of viral infection. However, whereas microbial pathogens can be recognized as nonself by immune receptors, misfolded protein assemblies evade detection, as they are host-derived. Here, we show that when misfolded tau assemblies enter the cell, they can be detected and neutralized via a danger response mediated by tau-associated antibodies and the cytosolic Fc receptor tripartite motif protein 21 (TRIM21). We developed fluorescent, morphology-based seeding assays that allow the formation of pathological tau aggregates to be measured in situ within 24 h in the presence of picomolar concentrations of tau seeds. We found that anti-tau antibodies accompany tau seeds into the cell, where they recruit TRIM21 shortly after entry. After binding, TRIM21 neutralizes tau seeds through the activity of the proteasome and the AAA ATPase p97/VCP in a similar manner to infectious viruses. These results establish that intracellular antiviral immunity can be redirected against host-origin endopathogens involved in neurodegeneration

The Role of Antibodies and Their Receptors in Protection Against Ordered Protein Assembly in Neurodegeneration.

Ordered assemblies of proteins are found in the postmortem brains of sufferers of several neurodegenerative diseases. The cytoplasmic microtubule associated protein tau and alpha-synuclein (αS) are found in an assembled state in Alzheimer's disease and Parkinson's disease, respectively. An accumulating body of evidence suggests a "prion-like" mechanism of spread of these assemblies through the diseased brain. Under this hypothesis, assembled variants of these proteins promote the conversion of native proteins to the assembled state. This likely inflicts pathology on cells of the brain through a toxic gain-of-function mechanism. Experiments in animal models of tau and αS pathology have demonstrated that the passive transfer of anti-tau or anti-αS antibodies induces a reduction in the levels of assembled proteins. This is further accompanied by improvements in neurological function and preservation of brain volume. Immunotherapy is therefore considered one of the brightest hopes as a therapeutic avenue in an area currently without disease-modifying therapy. Following a series of disappointing clinical trials targeting beta-amyloid, a peptide that accumulates in the extracellular spaces of the AD brain, attention is turning to active and passive immunotherapies that target tau and αS. However, there are several remaining uncertainties concerning the mechanism by which antibodies afford protection against self-propagating protein conformations. This review will discuss current understanding of how antibodies and their receptors can be brought to bear on proteins involved in neurodegeneration. Parallels will be made to antibody-mediated protection against classical viral infections. Common mechanisms that may contribute to protection against self-propagating protein conformations include blocking the entry of protein "seeds" to cells, clearance of immune complexes by microglia, and the intracellular protein degradation pathway initiated by cytoplasmic antibodies via the Fc receptor TRIM21. As with anti-viral immunity, protective mechanisms may be accompanied by the activation of immune signaling pathways and we will discuss the suitability of such activation in the neurological setting