12,560 research outputs found

    Interactions between Dpr11 and DIP-γ control selection of amacrine neurons in Drosophila color vision circuits

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    Drosophila R7 UV photoreceptors (PRs) are divided into yellow (y) and pale (p) subtypes. yR7 PRs express the Dpr11 cell surface protein and are presynaptic to Dm8 amacrine neurons (yDm8) that express Dpr11’s binding partner DIP-γ, while pR7 PRs synapse onto DIP-γ-negative pDm8. Dpr11 and DIP-γ expression patterns define ‘yellow’ and ‘pale’ color vision circuits. We examined Dm8 neurons in these circuits by electron microscopic reconstruction and expansion microscopy. DIP-γ and dpr11 mutations affect the morphologies of yDm8 distal (‘home column’) dendrites. yDm8 neurons are generated in excess during development and compete for presynaptic yR7 PRs, and interactions between Dpr11 and DIP-γ are required for yDm8 survival. These interactions also allow yDm8 neurons to select yR7 PRs as their appropriate home column partners. yDm8 and pDm8 neurons do not normally compete for survival signals or R7 partners, but can be forced to do so by manipulation of R7 subtype fate

    Subtle pH differences trigger single residue motions for moderating conformations of calmodulin

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    This study reveals the essence of ligand recognition mechanisms by which calmodulin (CaM) controls a variety of Ca2+ signaling processes. We study eight forms of calcium-loaded CaM each with distinct conformational states. Reducing the structure to two degrees of freedom conveniently describes main features of the conformational changes of CaM via simultaneous twist-bend motions of the two lobes. We utilize perturbation-response scanning (PRS) technique, coupled with molecular dynamics simulations. PRS is based on linear response theory, comprising sequential application of directed forces on selected residues followed by recording the resulting protein coordinates. We analyze directional preferences of the perturbations and resulting conformational changes. Manipulation of a single residue reproduces the structural change more effectively than that of single/pairs/triplets of collective modes of motion. Our findings also give information on how the flexible linker acts as a transducer of binding information to distant parts of the protein. Furthermore, by perturbing residue E31 located in one of the EF hand motifs in a specific direction, it is possible to induce conformational change relevant to five target structures. Independently, using four different pKa calculation strategies, we find this particular residue to be the charged residue (out of a total of 52), whose ionization state is most sensitive to subtle pH variations in the physiological range. It is plausible that at relatively low pH, CaM structure is less flexible. By gaining charged states at specific sites at a pH value around 7, such as E31 found in the present study, local conformational changes in the protein will lead to shifts in the energy landscape, paving the way to other conformational states. These findings are in accordance with Fluorescence Resonance Energy Transfer (FRET) measured shifts in conformational distributions towards more compact forms with decreased pH. They also corroborate mutational studies and proteolysis results which point to the significant role of E31 in CaM dynamics

    CD4+ T-cell responses to Epstein-Barr virus (EBV) latent-cycle antigens and the recognition of EBV-transformed lymphoblastoid cell lines

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    There is considerable interest in the potential of Epstein-Barr virus (EBV) latent antigen-specific CD4+^+ T cells to act as direct effectors controlling EBV-induced B lymphoproliferations. Such activity would require direct CD4+^+ T-cell recognition of latently infected cells through epitopes derived from endogenously expressed viral proteins and presented on the target cell surface in association with HLA class II molecules. It is therefore important to know how often these conditions are met. Here we provide CD4+^+ epitope maps for four EBV nuclear antigens, EBNA1, -2, -3A, and -3C, and establish CD4+^+ T-cell clones against 12 representative epitopes. For each epitope we identify the relevant HLA class II restricting allele and determine the efficiency with which epitope-specific effectors recognize the autologous EBV-transformed B-lymphoblastoid cell line (LCL). The level of recognition measured by gamma interferon release was consistent among clones to the same epitope but varied between epitopes, with values ranging from 0 to 35% of the maximum seen against the epitope peptide-loaded LCL. These epitope-specific differences, also apparent in short-term cytotoxicity and longer-term outgrowth assays on LCL targets, did not relate to the identity of the source antigen and could not be explained by the different functional avidities of the CD4+^+ clones; rather, they appeared to reflect different levels of epitope display at the LCL surface. Thus, while CD4+^+ T-cell responses are detectable against many epitopes in EBV latent proteins, only a minority of these responses are likely to have therapeutic potential as effectors directly recognizing latently infected target cells

    In silico prediction of mutant HIV-1 proteases cleaving a target sequence

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    HIV-1 protease represents an appealing system for directed enzyme re-design, since it has various different endogenous targets, a relatively simple structure and it is well studied. Recently Chaudhury and Gray (Structure (2009) 17: 1636 -- 1648) published a computational algorithm to discern the specificity determining residues of HIV-1 protease. In this paper we present two computational tools aimed at re-designing HIV-1 protease, derived from the algorithm of Chaudhuri and Gray. First, we present an energy-only based methodology to discriminate cleavable and non cleavable peptides for HIV-1 proteases, both wild type and mutant. Secondly, we show an algorithm we developed to predict mutant HIV-1 proteases capable of cleaving a new target substrate peptide, different from the natural targets of HIV-1 protease. The obtained in silico mutant enzymes were analyzed in terms of cleavability and specificity towards the target peptide using the energy-only methodology. We found two mutant proteases as best candidates for specificity and cleavability towards the target sequence

    Metabotropic glutamate 2/3 receptors and epigenetic modifications in psychotic disorders: a review

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    Schizophrenia and Bipolar Disorder are chronic psychiatric disorders, both considered as "major psychosis"; they are thought to share some pathogenetic factors involving a dysfunctional gene x environment interaction. Alterations in the glutamatergic transmission have been suggested to be involved in the pathogenesis of psychosis. Our group developed an epigenetic model of schizophrenia originated by Prenatal Restraint Stress (PRS) paradigm in mice. PRS mice developed some behavioral alterations observed in schizophrenic patients and classic animal models of schizophrenia, i.e. deficits in social interaction, locomotor activity and prepulse inhibition. They also showed specific changes in promoter DNA methylation activity of genes related to schizophrenia such as reelin, BDNF and GAD67, and altered expression and function of mGlu2/3 receptors in the frontal cortex. Interestingly, behavioral and molecular alterations were reversed by treatment with mGlu2/3 agonists. Based on these findings, we speculate that pharmacological modulation of these receptors could have a great impact on early phase treatment of psychosis together with the possibility to modulate specific epigenetic key protein involved in the development of psychosis. In this review, we will discuss in more details the specific features of the PRS mice as a suitable epigenetic model for major psychosis. We will then focus on key proteins of chromatin remodeling machinery as potential target for new pharmacological treatment through the activation of metabotropic glutamate receptors

    Cadmium and arsenic-induced-stress differentially modulates Arabidopsis root architecture, peroxisome distribution, enzymatic activities and their nitric oxide content

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    In plant cells, cadmium (Cd) and arsenic (As) exert toxicity mainly by inducing oxidative stress through an imbalance between the production of reactive oxygen species (ROS) and reactive nitrogen species (RNS), and their detoxification. Nitric oxide (NO) is a RNS acting as signalling molecule coordinating plant development and stress responses, but also as oxidative stress inducer, depending on its cellular concentration. Peroxisomes are versatile organelles involved in plant metabolism and signalling, with a role in cellular redox balance thanks to their antioxidant enzymes, and their RNS (mainly NO) and ROS. This study analysed Cd or As effects on peroxisomes, and NO production and distribution in the root system, including primary root (PR) and lateral roots (LRs). Arabidopsis thaliana wild-type and transgenic plants enabling peroxisomes to be visualized in vivo, through the expression of the 35S-cyan fluorescent protein fused to the peroxisomal targeting signal1 (PTS1) were used. Peroxisomal enzymatic activities including the antioxidant catalase, the H2O2-generating glycolate oxidase, and the hydroxypyruvate reductase, and root system morphology were also evaluated under Cd/As exposure. Results showed that Cd and As differently modulate these activities, however, catalase activity was inhibited by both. Moreover, Arabidopsis root system was altered, with the pollutants differently affecting PR growth, but similarly enhancing LR formation. Only in the PR apex, and not in LR one, Cd more than As caused significant changes in peroxisome distribution, size, and in peroxisomal NO content. By contrast, neither pollutant caused significant changes in peroxisomes size and peroxisomal NO content in the LR apex
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