The rockefeller university

Glial cells surround neuronal endings to create enclosed compartments required for neuronal function. This architecture is seen at excitatory synapses and at sensory neuron receptive endings. Despite the prevalence and importance of these compartments, how they form is not known. We used the main sensory organ of C. elegans, the amphid, to investigate this issue. daf-6/Patched-related is a glia-expressed gene previously implicated in amphid sensory compartment morphogenesis. By comparing time series of electron-microscopy (EM) reconstructions of wild-type and daf-6 mutant embryos, we show that daf-6 acts to restrict compartment size. From a genetic screen, we found that mutations in the gene lit-1/Nemo-like kinase (NLK) suppress daf-6. EM and genetic studies demonstrate that lit-1 acts within glia, in counterbalance to daf-6, to promote sensory compartment expansion. Although LIT-1 has been shown to regulate Wnt signaling, our genetic studies demonstrate a novel, Wnt-independent role for LIT-1 in sensory compartment size control. The LIT-1 activator MOM-4/TAK1 is also important for compartment morphogenesis and both proteins line the glial sensory compartment. LIT-1 compartment localization is important for its function and requires neuronal signals. Furthermore, the conserved LIT-1 C-terminus is necessary and sufficient for this localization. Two-hybrid and co-immunoprecipitation studies demonstrate that the LIT-1 C-terminus binds both actin and the Wiskott-Aldrich syndrome protein (WASP), an actin regulator. We use fluorescence light microscopy and fluorescence EM methodology to show that actin is highly enriched around the amphid sensory compartment. Finally, our genetic studies demonstrate that WASP is important for compartment expansion and functions in the same pathway as LIT-1. The studies presented here uncover a novel, Wnt-independent role for the conserved Nemo-like kinase LIT-1 in controlling cell morphogenesis in conjunction with the actin cytoskeleton. Our results suggest that the opposing daf-6 and lit-1 glial pathways act together to control sensory compartment size.

The axons of C. elegans left and right AWC olfactory neurons communicate at synapses through a calcium-signaling complex to regulate stochastic asymmetric cell identities called AWC(ON) and AWC(OFF). However, it is not known how the calcium-signaling complex, which consists of UNC-43/CaMKII, TIR-1/SARM adaptor protein and NSY-1/ASK1 MAPKKK, is localized to postsynaptic sites in the AWC axons for this lateral interaction. Here, we show that microtubule-based localization of the TIR-1 signaling complex to the synapses regulates AWC asymmetry. Similar to unc-43, tir-1 and nsy-1 loss-of-function mutants, specific disruption of microtubules in AWC by nocodazole generates two AWC(ON) neurons. Reduced localization of UNC-43, TIR-1 and NSY-1 proteins in the AWC axons strongly correlates with the 2AWC(ON) phenotype in nocodazole-treated animals. We identified kinesin motor unc-104/kif1a mutants for enhancement of the 2AWC(ON) phenotype of a hypomorphic tir-1 mutant. Mutations in unc-104, like microtubule depolymerization, lead to a reduced level of UNC-43, TIR-1 and NSY-1 proteins in the AWC axons. In addition, dynamic transport of TIR-1 in the AWC axons is dependent on unc-104, the primary motor required for the transport of presynaptic vesicles. Furthermore, unc-104 acts non-cell autonomously in the AWC(ON) neuron to regulate the AWC(OFF) identity. Together, these results suggest a model in which UNC-104 may transport some unknown presynaptic factor(s) in the future AWC(ON) cell that non-cell autonomously control the trafficking of the TIR-1 signaling complex to postsynaptic regions of the AWC axons to regulate the AWC(OFF) identity.

A long-sought, and thus far elusive, goal has been to develop drugs to manage diseases of excitability. One such disease that affects millions each year is cardiac arrhythmia, which occurs when electrical impulses in the heart become disordered, sometimes causing sudden death. Pharmacological management of cardiac arrhythmia has failed because it is not possible to predict how drugs that target cardiac ion channels, and have intrinsically complex dynamic interactions with ion channels, will alter the emergent electrical behavior generated in the heart. Here, we applied a computational model, which was informed and validated by experimental data, that defined key measurable parameters necessary to simulate the interaction kinetics of the anti-arrhythmic drugs flecainide and lidocaine with cardiac sodium channels. We then used the model to predict the effects of these drugs on normal human ventricular cellular and tissue electrical activity in the setting of a common arrhythmia trigger, spontaneous ventricular ectopy. The model forecasts the clinically relevant concentrations at which flecainide and lidocaine exacerbate, rather than ameliorate, arrhythmia. Experiments in rabbit hearts and simulations in human ventricles based on magnetic resonance images validated the model predictions. This computational framework initiates the first steps toward development of a virtual drug-screening system that models drug-channel interactions and predicts the effects of drugs on emergent electrical activity in the heart.

The annulus is an electron-dense ring structure connecting the midpiece and the principal piece of the mammalian sperm flagellum. Proteins from the septin family have been shown to localize to the annulus. A septin complex is assembled early in spermiogenesis with the cochaperone DNAJB13 and, in mature sperm, associates with Testis Anion Transporter 1; SLC26A8 (Tat1), a transmembrane protein of the SLC26 family. Studies in mice have shown that the annulus acts as a barrier to protein diffusion and controls correct organization of the midpiece. Consistent with these findings, absence of the annulus is associated with flagellum differentiation defects and asthenozoospermia in humans.

A measurement is presented of the charged hadron multiplicity in hadronic PbPb collisions, as a function of pseudorapidity and centrality, at a collision energy of 2.76 TeV per nucleon pair. The data sample is collected using the CMS detector and a minimum-bias trigger, with the CMS solenoid off. The number of charged hadrons is measured both by counting the number of reconstructed particle hits and by forming hit doublets of pairs of layers in the pixel detector. The two methods give consistent results. The charged hadron multiplicity density, dN(ch)/d eta vertical bar(eta=0), for head-on collisions is found to be 1612 +/- 55, where the uncertainty is dominated by systematic effects. Comparisons of these results to previous measurements and to various models are also presented.

CD40, a member of the tumor necrosis factor receptor (TNFR) superfamily, is expressed on antigen-presenting cells (APCs) and is essential for immune activation. Although agonistic CD40 antibodies have been developed for immunotherapy, their clinical efficacy has been limited. We have found that coengagement of the Fc domain of agonistic CD40 monoclonal antibodies (mAbs) with the inhibitory Fc gamma receptor Fc gamma RIIB is required for immune activation. Direct comparison of mAbs to CD40 enhanced for activating Fc gamma R binding, hence capable of cytotoxicity, or for inhibitory Fc gamma RIIB binding, revealed that enhancing Fc gamma RIIB binding conferred immunostimulatory activity and considerably greater anti-tumor responses. This unexpected requirement for Fc gamma RIIB in enhancing CD40-mediated immune activation has direct implications for the design of agonistic antibodies to TNFR as therapeutics.

A 33-year-old man was admitted in hospital due to fever, generalized lymphadenopathy, and hepatosplenomegaly. He had a history of anti-tuberculosis treatment in the previous 3 years. Despite normal chest radiograph, a sputum sample was smear-positive for acid-fast bacilli, and polymerase chain reaction was positive for Mycobacterium tuberculosis complex. Drug susceptibility test revealed resistance to isoniazid and rifampin. Evaluation of the patient's immune system revealed IL-12R beta 1 deficiency. The patient died of disseminated tuberculosis (TB), despite appropriate antibiotic treatment. This is the first IL-12 receptor-deficient patient presenting with disseminated TB in adulthood, without any previous relevant medical history. This diagnosis should be considered in selected adult patients with unexplained, overwhelming TB. IL-12R beta 1 deficiency is a genetic etiology of severe TB in adults and should be considered in adult patients with disseminated TB.

A single gram of soil is predicted to contain thousands of unique bacterial species. The majority of these species remain recalcitrant to standard culture methods, prohibiting their use as sources of unique bioactive small molecules. The cloning and analysis of DNA extracted directly from environmental samples (environmental DNA, eDNA) provides a means of exploring the biosynthetic capacity of natural bacterial populations. Environmental DNA libraries contain large reservoirs of bacterial genetic diversity from which new secondary metabolite gene clusters can be systematically recovered and studied. The identification and heterologous expression of type II polyketide synthase-containing eDNA clones is reported here. Functional analysis of three soil DNA-derived polyketide synthase systems in Streptomyces albus revealed diverse metabolites belonging to well-known, rare, and previously uncharacterized structural families. The first of these systems is predicted to encode the production of the known antibiotic landomycin E. The second was found to encode the production of a metabolite with a previously uncharacterized pentacyclic ring system. The third was found to encode the production of unique KB-3346-5 derivatives, which show activity against methicillin-resistant Staphylococcus aureus and vancomycin-resistant Enterococcus faecalis. These results, together with those of other small-molecule-directed metagenomic studies, suggest that culture-independent approaches are capable of accessing biosynthetic diversity that has not yet been extensively explored using culture-based methods. The large-scale functional screening of eDNA clones should be a productive strategy for generating structurally previously uncharacterized chemical entities for use in future drug development efforts.

The observation of the bottom, strange baryon Xi(0)(b) through the decay chain Xi(0)(b) -> Xi(+)(c)pi(-), where Xi(+)(c) -> Xi(-) pi(+) pi(+), Xi(-) -> Lambda pi(-), and Lambda -> p pi(-), is reported by using data corresponding to an integrated luminosity of 4.2 fb(-1) from p (p) over bar collisions at root s = 1.96 TeV recorded with the Collider Detector at Fermilab. A signal of 25.3(-5.4)(+5.6) candidates is observed whose probability of arising from a background fluctuation is 3.6 x 10(-12), corresponding to 6.8 Gaussian standard deviations. The Xi(0)(b) mass is measured to be 5787.8 +/- 5.0(stat) +/- 1.3(syst) MeV/c(2). In addition, the Xi(-)(b) baryon is observed through the process Xi(-)(b) -> Xi(0)(c)pi(-), where Xi(0)(c) -> Xi(-) pi(+), Xi(-) -> Lambda pi(-), and Lambda -> p pi(-).

Obese, leptin deficient obob mice have profoundly decreased activity and increased food seeking behavior. The decreased activity has been attributed to obesity. In mice, we tested the hypothesis that leptin increases total locomotor activity but inhibits food anticipatory activity. We also sought to determine if leptin induced increases in total locomotor activity are independent of changes in body weight and obesity. We studied obob mice and also created a novel transgenic mouse where leptin is over-expressed in a tetracycline-off system and can be abruptly and non-invasively suppressed by doxycycline within few hours. The studies were performed using two independent behavioral assays: home cage activity (HCA) and running wheel activity (RWA). Systemic administration of leptin (150 ng/hr) to obob mice produced a 122% +/- 30% (mean +/- SEM) increase (p <= 0.01) in locomotor activity within 2 days In addition, cerebroventricular administration of leptin (5 ng/hr) also produced an early and progressive increase in total locomotor activity beginning on the 1st day (+28 +/- 8%; p <= 0.05) and increasing to +69 +/- 23% on day 3 without a decrease in body weight during this time. The increase in activity was restricted to the dark phase. Conversely, in a tet-off transgenic obob mouse line, acute leptin suppression reduced spontaneous locomotor activity. To further define activities that are leptin regulated, we assayed food anticipatory activity (FAA) and found that it was markedly augmented in obob mice compared to wild type mice (+38 +/- 6.7 in obob vs +20 +/- 6.3% in wild type at peak; mean +/- SEM; p <= 0.001) and abolished by leptin. Although melanocortin-3 receptors (MC3R) reportedly mediate FAA, we found augmented FAA and preserved inhibitory effects of leptin on FAA in MC3R-/-obob mice. In summary, this study demonstrates that total activity and FAA are regulated independently by leptin. Leptin, acting in the central nervous system and at physiologic levels, produces early increases in locomotor activity before substantial weight loss. In contrast, leptin suppresses augmented food anticipatory activity in obob mice.