200 research outputs found
Recommended from our members
Annual Report of the International Nuclear Research Initiative Osmose Project (FY05).
The goal of the OSMOSE program is to measure the reactivity effect of minor actinides in known neutron spectra of interest to the Generation-IV reactor program and other programs and to create a database of these results for use as an international benchmark for the minor actinides. The results are then compared to calculational models to verify and validate integral absorption cross-sections for the minor actinides. The OSMOSE program includes all aspects of the experimental program -- including the fabrication of fuel pellets and samples, the oscillation of the samples in the MINERVE reactor for the measurement of the reactivity effect, reactor physics modeling of the MINERVE reactor, and the data analysis and interpretation of the experimental results
FFAT motif phosphorylation controls formation and lipid transfer function of interâorganelle contacts
Organelles are physically connected in membrane contact sites. The endoplasmic reticulum possesses three major receptors, VAPâA, VAPâB, and MOSPD2, which interact with proteins at the surface of other organelles to build contacts. VAPâA, VAPâB, and MOSPD2 contain an MSP domain, which binds a motif named FFAT (two phenylalanines in an acidic tract). In this study, we identified a nonâconventional FFAT motif where a conserved acidic residue is replaced by a serine/threonine. We show that phosphorylation of this serine/threonine is critical for nonâconventional FFAT motifs (named PhosphoâFFAT) to be recognized by the MSP domain. Moreover, structural analyses of the MSP domain alone or in complex with conventional and PhosphoâFFAT peptides revealed new mechanisms of interaction. Based on these new insights, we produced a novel prediction algorithm, which expands the repertoire of candidate proteins with a PhosphoâFFAT that are able to create membrane contact sites. Using a prototypical tethering complex made by STARD3 and VAP, we showed that phosphorylation is instrumental for the formation of ERâendosome contacts, and their sterol transfer function. This study reveals that phosphorylation acts as a general switch for interâorganelle contacts
The structure of the scaffold nucleoporin Nup120 reveals a new and unexpected domain architecture
Nucleocytoplasmic transport is mediated by nuclear pore complexes (NPCs), enormous protein assemblies residing in circular openings in the nuclear envelope. The NPC is modular, with transient and stable components. The stable core is essentially built from two multiprotein complexes, the Y-shaped heptameric Nup84 complex and the Nic96 complex, arranged around an eightfold axis. We present the crystal structure of Nup120[subscript 1-757], one of the two short arms of the Y-shaped Nup84 complex. The protein adopts a compact oval shape built around a novel bipartite α-helical domain intimately integrated with a ÎČ-propeller domain. The domain arrangement is substantially different from the Nup85âąSeh1 complex, which forms the other short arm of the Y. With the data presented here, we establish that all three branches of the Y-shaped Nup84 complex are tightly connected by helical interactions and that the ÎČ-propellers likely form interaction site(s) to neighboring complexes.National Institutes of Health (U.S.) (Grant GM77537)Pew Charitable Trusts (Scholar Award
Non-Metabolic Membrane Tubulation and Permeability Induced by Bioactive Peptides
BACKGROUND: Basic cell-penetrating peptides are potential vectors for therapeutic molecules and display antimicrobial activity. The peptide-membrane contact is the first step of the sequential processes leading to peptide internalization and cell activity. However, the molecular mechanisms involved in peptide-membrane interaction are not well understood and are frequently controversial. Herein, we compared the membrane activities of six basic peptides with different size, charge density and amphipaticity: Two cell-penetrating peptides (penetratin and R9), three amphipathic peptides and the neuromodulator substance P. METHODOLOGY/PRINCIPAL FINDINGS: Experiments of X ray diffraction, video-microscopy of giant vesicles, fluorescence spectroscopy, turbidimetry and calcein leakage from large vesicles are reported. Permeability and toxicity experiments were performed on cultured cells. The peptides showed differences in bilayer thickness perturbations, vesicles aggregation and local bending properties which form lipidic tubular structures. These structures invade the vesicle lumen in the absence of exogenous energy. CONCLUSIONS/SIGNIFICANCE: We showed that the degree of membrane permeabilization with amphipathic peptides is dependent on both peptide size and hydrophobic nature of the residues. We propose a model for peptide-induced membrane perturbations that explains the differences in peptide membrane activities and suggests the existence of a facilitated âphysical endocytosis,â which represents a new pathway for peptide cellular internalization
Cross-Talk between Signaling Pathways Can Generate Robust Oscillations in Calcium and cAMP
BACKGROUND:To control and manipulate cellular signaling, we need to understand cellular strategies for information transfer, integration, and decision-making. A key feature of signal transduction is the generation of only a few intracellular messengers by many extracellular stimuli. METHODOLOGY/PRINCIPAL FINDINGS:Here we model molecular cross-talk between two classic second messengers, cyclic AMP (cAMP) and calcium, and show that the dynamical complexity of the response of both messengers increases substantially through their interaction. In our model of a non-excitable cell, both cAMP and calcium concentrations can oscillate. If mutually inhibitory, cross-talk between the two second messengers can increase the range of agonist concentrations for which oscillations occur. If mutually activating, cross-talk decreases the oscillation range, but can generate 'bursting' oscillations of calcium and may enable better filtering of noise. CONCLUSION:We postulate that this increased dynamical complexity allows the cell to encode more information, particularly if both second messengers encode signals. In their native environments, it is unlikely that cells are exposed to one stimulus at a time, and cross-talk may help generate sufficiently complex responses to allow the cell to discriminate between different combinations and concentrations of extracellular agonists
Phospholipase C-eta enzymes as putative protein kinase C and Ca2+ signalling components in neuronal and neuroendocrine tissues
Phosphoinositol-specific phospholipase C enzymes (PLCs) are central to inositol lipid signalling pathways, facilitating intracellular Ca2+ release and protein kinase C activation. A sixth class of phosphoinositol-specific PLC with a novel domain structure, PLC-eta (PLCeta) has recently been discovered in mammals. Recent research, reviewed here, shows that this class consists of two enzymes, PLCeta1 and PLCeta2. Both enzymes hydrolyze phosphatidylinositol 4,5-bisphosphate and are more sensitive to Ca2+ than other PLC isozymes and are likely to mediate G-protein-coupled receptor (GPCR) signalling pathways. Both enzymes are expressed in neuron-enriched regions, being abundant in the brain. We demonstrate that they are also expressed in neuroendocrine cell lines. PLCeta enzymes therefore represent novel proteins influencing intracellular Ca2+ dynamics and protein kinase C activation in the brain and neuroendocrine systems as putative mediation of GPCR regulation
CNS Delivery Via Adsorptive Transcytosis
Adsorptive-mediated transcytosis (AMT) provides a means for brain delivery of medicines across the blood-brain barrier (BBB). The BBB is readily equipped for the AMT process: it provides both the potential for binding and uptake of cationic molecules to the luminal surface of endothelial cells, and then for exocytosis at the abluminal surface. The transcytotic pathways present at the BBB and its morphological and enzymatic properties provide the means for movement of the molecules through the endothelial cytoplasm. AMT-based drug delivery to the brain was performed using cationic proteins and cell-penetrating peptides (CPPs). Protein cationization using either synthetic or natural polyamines is discussed and some examples of diamine/polyamine modified proteins that cross BBB are described. Two main families of CPPs belonging to the Tat-derived peptides and Syn-B vectors have been extensively used in CPP vector-mediated strategies allowing delivery of a large variety of small molecules as well as proteins across cell membranes in vitro and the BBB in vivo. CPP strategy suffers from several limitations such as toxicity and immunogenicityâlike the cationization strategyâas well as the instability of peptide vectors in biological media. The review concludes by stressing the need to improve the understanding of AMT mechanisms at BBB and the effectiveness of cationized proteins and CPP-vectorized proteins as neurotherapeutics
Molecular architecture of the Nup84âNup145CâSec13 edge element in the nuclear pore complex lattice
Nuclear pore complexes (NPCs) facilitate all nucleocytoplasmic transport. These massive protein assemblies are modular, with a stable structural scaffold supporting more dynamically attached components. The scaffold is made from multiple copies of the heptameric Y complex and the heteromeric Nic96 complex. We previously showed that members of these core subcomplexes specifically share an ACE1 fold with Sec31 of the COPII vesicle coat, and we proposed a lattice model for the NPC based on this commonality. Here we present the crystal structure of the heterotrimeric 134-kDa complex of Nup84âNup145CâSec13 of the Y complex. The heterotypic ACE1 interaction of Nup84 and Nup145C is analogous to the homotypic ACE1 interaction of Sec31 that forms COPII lattice edge elements and is inconsistent with the alternative 'fence-like' NPC model. We construct a molecular model of the Y complex and compare the architectural principles of COPII and NPC lattices.National Institutes of Health (U.S.) (Grant GM77537)Pew Charitable Trusts (Scholar Award
- âŠ