G Protein Subunit Dissociation and Translocation Regulate Cellular Response to Receptor Stimulation

We examined the role of G proteins in modulating the response of living cells to receptor activation. The response of an effector, phospholipase C-β to M3 muscarinic receptor activation was measured using sensors that detect the generation of inositol triphosphate or diacylglycerol. The recently discovered translocation of Gβγ from plasma membrane to endomembranes on receptor activation attenuated this response. A FRET based G protein sensor suggested that in contrast to translocating Gβγ, non-translocating Gβγ subunits do not dissociate from the αq subunit on receptor activation leading to prolonged retention of the heterotrimer state and an accentuated response. M3 receptors with tethered αq induced differential responses to receptor activation in cells with or without an endogenous translocation capable γ subunit. G protein heterotrimer dissociation and βγ translocation are thus unanticipated modulators of the intensity of a cell's response to an extracellular signal

The electric vehicle in smart homes: a review and future perspectives

The electric mobility dissemination is forcing the adoption of new technologies and operation paradigms, not only focusing on smart grids, but also on smart homes. In fact, the emerging technologies for smart homes are also altering the conventional grids toward smart grids. By combining the key pillars of electric mobility and smart homes, this paper characterizes the paradigms of the electric vehicle (EV) in smart homes, presenting a review about the state of the art and establishing a relation with future perspectives. Since the smart home must be prepared to deal with the necessities of the EV, the analysis of both on board and off board battery charging systems are considered in the paper. Moreover, the in-clusion of renewable energy sources, energy storage systems, and dc electrical appliances in smart homes towards sustainability is also considered in this paper, but framed in the perspective of an EV off board battery charging system. As a pertinent contribution, this paper offers future perspectives for the EV in smart homes, including the possibility of ac, dc, and hybrid smart homes. Covering all of these aspects, exemplificative and key results are presented based on numerical simulations and experimental results obtained with a proof of concept prototype.FCT – Fundação para a Ciência e Tecnologia within the Project Scope: UID/CEC/00319/2019. This work has been supported by the FCT Project newERA4GRIDs PTDC/EEI-EEE/30283/2017, and by the FCT Project DAIPESEV PTDC/EEI-EEE/30382/2017. Tiago Sousa is supported by the doctoral scholarship SFRH/BD/134353/2017 granted by FC

Dynamics of Mechanical Signal Transmission through Prestressed Stress Fibers

Transmission of mechanical stimuli through the actin cytoskeleton has been proposed as a mechanism for rapid long-distance mechanotransduction in cells; however, a quantitative understanding of the dynamics of this transmission and the physical factors governing it remains lacking. Two key features of the actin cytoskeleton are its viscoelastic nature and the presence of prestress due to actomyosin motor activity. We develop a model of mechanical signal transmission through prestressed viscoelastic actin stress fibers that directly connect the cell surface to the nucleus. The analysis considers both temporally stationary and oscillatory mechanical signals and accounts for cytosolic drag on the stress fibers. To elucidate the physical parameters that govern mechanical signal transmission, we initially focus on the highly simplified case of a single stress fiber. The results demonstrate that the dynamics of mechanical signal transmission depend on whether the applied force leads to transverse or axial motion of the stress fiber. For transverse motion, mechanical signal transmission is dominated by prestress while fiber elasticity has a negligible effect. Conversely, signal transmission for axial motion is mediated uniquely by elasticity due to the absence of a prestress restoring force. Mechanical signal transmission is significantly delayed by stress fiber material viscosity, while cytosolic damping becomes important only for longer stress fibers. Only transverse motion yields the rapid and long-distance mechanical signal transmission dynamics observed experimentally. For simple networks of stress fibers, mechanical signals are transmitted rapidly to the nucleus when the fibers are oriented largely orthogonal to the applied force, whereas the presence of fibers parallel to the applied force slows down mechanical signal transmission significantly. The present results suggest that cytoskeletal prestress mediates rapid mechanical signal transmission and allows temporally oscillatory signals in the physiological frequency range to travel a long distance without significant decay due to material viscosity and/or cytosolic drag

Generation of recombinant hyperimmune globulins from diverse B-cell repertoires

Plasma-derived polyclonal antibody therapeutics, such as intravenous immunoglobulin, have multiple drawbacks, including low potency, impurities, insufficient supply and batch-to-batch variation. Here we describe a microfluidics and molecular genomics strategy for capturing diverse mammalian antibody repertoires to create recombinant multivalent hyperimmune globulins. Our method generates of diverse mixtures of thousands of recombinant antibodies, enriched for specificity and activity against therapeutic targets. Each hyperimmune globulin product comprised thousands to tens of thousands of antibodies derived from convalescent or vaccinated human donors or from immunized mice. Using this approach, we generated hyperimmune globulins with potent neutralizing activity against severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) in under 3 months, Fc-engineered hyperimmune globulins specific for Zika virus that lacked antibody-dependent enhancement of disease, and hyperimmune globulins specific for lung pathogens present in patients with primary immune deficiency. To address the limitations of rabbit-derived anti-thymocyte globulin, we generated a recombinant human version and demonstrated its efficacy in mice against graft-versus-host disease

The Human Serum Metabolome

Continuing improvements in analytical technology along with an increased interest in performing comprehensive, quantitative metabolic profiling, is leading to increased interest pressures within the metabolomics community to develop centralized metabolite reference resources for certain clinically important biofluids, such as cerebrospinal fluid, urine and blood. As part of an ongoing effort to systematically characterize the human metabolome through the Human Metabolome Project, we have undertaken the task of characterizing the human serum metabolome. In doing so, we have combined targeted and non-targeted NMR, GC-MS and LC-MS methods with computer-aided literature mining to identify and quantify a comprehensive, if not absolutely complete, set of metabolites commonly detected and quantified (with today's technology) in the human serum metabolome. Our use of multiple metabolomics platforms and technologies allowed us to substantially enhance the level of metabolome coverage while critically assessing the relative strengths and weaknesses of these platforms or technologies. Tables containing the complete set of 4229 confirmed and highly probable human serum compounds, their concentrations, related literature references and links to their known disease associations are freely available at http://www.serummetabolome.ca

The role of off-board EV battery chargers in smart homes and smart grids: operation with renewables and energy storage systems

Concerns about climate changes and environmental air pollution are leading to the adoption of new technologies for transportation, mainly based on vehicle electrification and the interaction with smart grids, and also with the introduction of renewable energy sources (RES) accompanied by energy storage systems (ESS). For these three fundamental pillars, new power electronics technologies are emerging to transform the electrical power grid, targeting a flexible and collaborative operation. As a distinctive factor, the vehicle electrification has stimulated the presence of new technologies in terms of power management, both for smart homes and smart grids. As the title indicates, this book chapter focuses on the role of off-board EV battery chargers in terms of operation modes and contextualization for smart homes and smart grids in terms of opportunities. Based on a review of on-board and off-board EV battery charging systems (EV-BCS), this chapter focus on the off-board EV-BCS framed with RES and ESS as a dominant system in future smart homes. Contextualizing these aspects, three distinct cases are considered: (1) An ac smart home using separate power converters, according to the considered technologies; (2) A hybrid ac and dc smart home with an off-board EV-BCS interfacing RES and ESS, and with the electrical appliances plugged-in to the ac power grid; (3) A dc smart home using a unified 2 off-board EV-BCS with a single interface for the electrical power grid, and with multiple dc interfaces (RES, ESS, and electrical appliances). The results for each case are obtained in terms of efficiency and power quality, demonstrating that the off-board EV-BCS, as a unified structure for smart homes, presents better results. Besides, the off-board EV-BCS can also be used as an important asset for the smart grid, even when the EV is not plugged-in at the smart home.(undefined

Evaluation of Greenbug and Yellow Sugarcane Aphid Feeding Behavior on Resistant and Susceptible Switchgrass Cultivars

Switchgrass (Panicum virgatum L.) is an emerging biofuel crop that serves as host for aphids. To discern the effects of plant age and possible resistance mechanisms, the feeding behavior of greenbugs (Schizaphis graminum Rondani.) and the yellow sugarcane aphid (Sipha flava Forbes.) was monitored on three diverse switchgrasses by the electrical penetration graph (EPG) technique. Callose deposition and genes associated with callose metabolism were also analyzed to discern their association with plant resistance. There was a strong host effect on greenbugs feeding on lowland cultivar Kanlow at the V3 stage of development, as compared to the greenbug-susceptible upland cultivar Summer and plants derived from Kanlow (♂) × Summer (♀) (K×S) crosses. These data confirmed that Kanlow at the V3 stage had antibiosis to greenbugs, which was absent in the Summer and K×S plants. In contrast, similar effects were not observed for yellow sugarcane aphids, excluding significant differences in the time to first probe on Kanlow plants at the V1 stage and reduction in time spent on pathway processes on Kanlow plants at the V3 stage. These data demonstrated that Kanlow plants may have multiple sources of resistance to the two aphids, and possibly some were phloem based. Microscopy of leaf sections stained with aniline blue for callose was suggestive of increased callose deposition in the sieve elements in Kanlow plants relative to Summer and K×S plants. RT-qPCR analysis of several genes associated with callose metabolism in infested plants was equivocal. Overall, these studies suggest the presence of multiple defense mechanisms against aphids in Kanlow plants, relative to Summer and K×S plants

Inositol 1,4,5- Trisphosphate Receptor Function in Drosophila Insulin Producing Cells

The Inositol 1,4,5- trisphosphate receptor (InsP3R) is an intracellular ligand gated channel that releases calcium from intracellular stores in response to extracellular signals. To identify and understand physiological processes and behavior that depends on the InsP3 signaling pathway at a systemic level, we are studying Drosophila mutants for the InsP3R (itpr) gene. Here, we show that growth defects precede larval lethality and both are a consequence of the inability to feed normally. Moreover, restoring InsP3R function in insulin producing cells (IPCs) in the larval brain rescues the feeding deficit, growth and lethality in the itpr mutants to a significant extent. We have previously demonstrated a critical requirement for InsP3R activity in neuronal cells, specifically in aminergic interneurons, for larval viability. Processes from the IPCs and aminergic domain are closely apposed in the third instar larval brain with no visible cellular overlap. Ubiquitous depletion of itpr by dsRNA results in feeding deficits leading to larval lethality similar to the itpr mutant phenotype. However, when itpr is depleted specifically in IPCs or aminergic neurons, the larvae are viable. These data support a model where InsP3R activity in non-overlapping neuronal domains independently rescues larval itpr phenotypes by non-cell autonomous mechanisms

Murine Leukemia Virus Spreading in Mice Impaired in the Biogenesis of Secretory Lysosomes and Ca2+-Regulated Exocytosis

Retroviruses have been observed to bud intracellularly into multivesicular bodies (MVB), in addition to the plasma membrane. Release from MVB is thought to occur by Ca(2+)-regulated fusion with the plasma membrane.To address the role of the MVB pathway in replication of the murine leukemia virus (MLV) we took advantage of mouse models for the Hermansky-Pudlak syndrome (HPS) and Griscelli syndrome. In humans, these disorders are characterized by hypopigmentation and immunological alterations that are caused by defects in the biogenesis and trafficking of MVBs and other lysosome related organelles. Neonatal mice for these disease models lacking functional AP-3, Rab27A and BLOC factors were infected with Moloney MLV and the spread of virus into bone marrow, spleen and thymus was monitored. We found a moderate reduction in MLV infection levels in most mutant mice, which differed by less than two-fold compared to wild-type mice. In vitro, MLV release form bone-marrow derived macrophages was slightly enhanced. Finally, we found no evidence for a Ca(2+)-regulated release pathway in vitro. Furthermore, MLV replication was only moderately affected in mice lacking Synaptotagmin VII, a Ca(2+)-sensor regulating lysosome fusion with the plasma membrane.Given that MLV spreading in mice depends on multiple rounds of replication even moderate reduction of virus release at the cellular level would accumulate and lead to a significant effect over time. Thus our in vivo and in vitro data collectively argue against an essential role for a MVB- and secretory lysosome-mediated pathway in the egress of MLV