Sequential forward and reverse transport of the Na+ Ca2+ exchanger generates Ca2+ oscillations within mitochondria

Mitochondrial Ca2+ homoeostasis regulates aerobic metabolism and cell survival. Ca2+ flux into mitochondria is mediated by the mitochondrial calcium uniporter (MCU) channel whereas Ca2+ export is often through an electrogenic Na+–Ca2+ exchanger. Here, we report remarkable functional versatility in mitochondrial Na+–Ca2+ exchange under conditions where mitochondria are depolarised. Following physiological stimulation of cell-surface receptors, mitochondrial Na+–Ca2+ exchange initially operates in reverse mode, transporting cytosolic Ca2+ into the matrix. As matrix Ca2+ rises, the exchanger reverts to its forward mode state, extruding Ca2+. Transitions between reverse and forward modes generate repetitive oscillations in matrix Ca2+. We further show that reverse mode Na+–Ca2+ activity is regulated by the mitochondrial fusion protein mitofusin 2. Our results demonstrate that reversible switching between transport modes of an ion exchanger molecule generates functionally relevant oscillations in the levels of the universal Ca2+ messenger within an organelle

Control of NFAT Isoform Activation and NFAT-Dependent Gene Expression through Two Coincident and Spatially Segregated Intracellular Ca 2+ Signals

© 2016 The Author(s) Excitation-transcription coupling, linking stimulation at the cell surface to changes in nuclear gene expression, is conserved throughout eukaryotes. How closely related coexpressed transcription factors are differentially activated remains unclear. Here, we show that two Ca2+-dependent transcription factor isoforms, NFAT1 and NFAT4, require distinct sub-cellular InsP3 and Ca2+ signals for physiologically sustained activation. NFAT1 is stimulated by sub-plasmalemmal Ca2+ microdomains, whereas NFAT4 additionally requires Ca2+ mobilization from the inner nuclear envelope by nuclear InsP3 receptors. NFAT1 is rephosphorylated (deactivated) more slowly than NFAT4 in both cytoplasm and nucleus, enabling a more prolonged activation phase. Oscillations in cytoplasmic Ca2+, long considered the physiological form of Ca2+ signaling, play no role in activating either NFAT protein. Instead, effective sustained physiological activation of NFAT4 is tightly linked to oscillations in nuclear Ca2+. Our results show how gene expression can be controlled by coincident yet geographically distinct Ca2+ signals, generated by a freely diffusible InsP3 message

Seasonal Prediction of Arabian Sea Marine Heatwaves

Marine heatwaves are known to have a detrimental impact on marine ecosystems, yet predicting when and where they will occur remains a challenge. Here, using a large ensemble of initialized predictions from an Earth System Model, we demonstrate skill in predictions of summer marine heatwaves over large marine ecosystems in the Arabian Sea seven months ahead. Retrospective forecasts of summer (June to August) marine heatwaves initialized in the preceding winter (November) outperform predictions based on observed frequencies. These predictions benefit from initialization during winters of medium to strong El Niño conditions, which have an impact on marine heatwave characteristics in the Arabian Sea. Our probabilistic predictions target spatial characteristics of marine heatwaves that are specifically useful for fisheries management, as we demonstrate using an example of Indian oil sardine (Sardinella longiceps)

Nuanced Interactions between AKAP79 and STIM1 with Orai1 Ca 2+ Channels at Endoplasmic Reticulum-Plasma Membrane Junctions Sustain NFAT Activation

A-kinase anchoring protein 79 (AKAP79) is a human scaffolding protein that organizes Ca2+/calmodulin-dependent protein phosphatase calcineurin, calmodulin, cAMP-dependent protein kinase, protein kinase C, and the transcription factor nuclear factor of activated T cells (NFAT1) into a signalosome at the plasma membrane. Upon Ca2+ store depletion, AKAP79 interacts with the N-terminus of STIM1-gated Orai1 Ca2+ channels, enabling Ca2+ nanodomains to stimulate calcineurin. Calcineurin then dephosphorylates and activates NFAT1, which then translocates to the nucleus. A fundamental question is how signalosomes maintain long-term signaling when key effectors are released and therefore removed beyond the reach of the activating signal. Here, we show that the AKAP79-Orai1 interaction is considerably more transient than that of STIM1-Orai1. Free AKAP79, with calcineurin and NFAT1 in tow, is able to replace rapidly AKAP79 devoid of NFAT1 on Orai1, in the presence of continuous Ca2+ entry. We also show that Ca2+ nanodomains near Orai1 channels activate almost the entire cytosolic pool of NFAT1. Recycling of inactive NFAT1 from the cytoplasm to AKAP79 in the plasma membrane, coupled with the relatively weak interaction between AKAP79 and Orai1, maintain excitation-transcription coupling. By measuring rates for AKAP79-NFAT interaction, we formulate a mathematical model that simulates NFAT dynamics at the plasma membrane

Conformational surveillance of Orai1 by a rhomboid intramembrane protease prevents inappropriate CRAC channel activation

Calcium influx through plasma membrane calcium release-activated calcium (CRAC) channels, which are formed of hexamers of Orai1, is a potent trigger for many important biological processes, most notably in T cell-mediated immunity. Through a bioinformatics-led cell biological screen, we have identified Orai1 as a substrate for the rhomboid intramembrane protease RHBDL2. We show that RHBDL2 prevents stochastic calcium signaling in unstimulated cells through conformational surveillance and cleavage of inappropriately activated Orai1. A conserved disease-linked proline residue is responsible for RHBDL2’s recognizing the active conformation of Orai1, which is required to sharpen switch-like signaling triggered by store-operated calcium entry. Loss of RHBDL2 control of CRAC channel activity causes severe dysregulation of downstream CRAC channel effectors, including transcription factor activation, inflammatory cytokine expression, and T cell activation. We propose that this surveillance function may represent an ancient activity of rhomboid proteases in degrading unwanted signaling proteins

Ancient Egyptian soup for treating asthma: Cox and Intal

Chromone derivatives exhibiting unusual biological activity have been studied in these laboratories for several years. One of these, disodium cromoglycate (FLP670), has been found to inhibit specifically the liberation of the mediators of anaphylaxis initiated by the interaction of antigen with regain type antibodies. It is neither a bronchodilator nor an antiinflammatory agent and its action is distinct from that of corticosteroids. In experiments on an asthmatic volunteer, inhaling the compound inhibited the acute effect of subsequent antigen inhalation. In patients suffering from allergic asthma, inhalation of this compound was followed by clinical improvement

Neuronal Ca 2+ signalling: pathways and targets

Imaging and models of brain function. Held at Heidelberg, Germany. 15–17, March, 2001

Calcium signalling in immune cells

Inappropriate stimulation of mast cells can trigger allergies including asthma, allergic rhinitis and eczema which, combined, affect almost 30% of the population in western societies. Mast cell activation begins with aggregation of IgE receptors in response to antigen. This then triggers a series of reactions resulting in the tyrosine phosphorylation of Syk kinase, PKC activation and ultimately both degranulation and secretion of leukotrienes and cytokines. CRAC channels are expressed on mast cells, and are essential for IgE-mediated mast cell activation. Previous work in our laboratory has shown that local Ca2+ influx through CRAC channels activates Ca2+-dependent phopholipase A₂, ERK and 5-lipoxygenase, resulting in LTC₄ secretion from mast cells. Therefore, I have investigated how Ca2+ microdomains through CRAC channels are detected and how they trigger cellular responses. I find that phosphorylation of Syk following antigen stimulation is enhanced by Ca2+ influx through CRAC channels. I also show synergy between CRAC channels and antigen in activating Syk. These findings reveal a novel positive feedback step in mast cell activation, where local Ca2+ entry through CRAC channels activates Syk which, in turn, supports CRAC channels. Earlier work from our group has demonstrated that in RBL cells, Ca2+ influx through CRAC channels induces expression of the gene c-fos, an important regulator of pro-inflammatory gene expression. I have discovered that local Ca2+ entry is sensed by the non-receptor tyrosine kinase Syk, which accumulates at the cell periphery. Syk then signals to the nucleus through recruitment of the transcription factor STAT5. The results therefore identify Syk as a new link in excitation-transcription coupling, converting local Ca2+ influx into expression of genes that are essential for immune cell activation. Activation of G protein-coupled cysteinyl leukotriene type I receptors by the pro-inflammatory molecule LTC₄ is tightly linked to immune cell function and the receptor is an established therapeutic target for allergies including asthma. Desensitization of cysteinyl leukotriene type I receptors arises following protein kinase C-dependent phosphorylation of three serine residues in the receptor C-terminus. Here I show that abolishing leukotriene receptor desensitization suppresses agonist-driven gene expression. Physiological concentrations of LTC₄ led to repetitive cytoplasmic Ca2+ oscillations, which were accompanied by the opening of store-operated CRAC channels in the plasma membrane. Ca2+ microdomains near the open channels were relayed to the nucleus to increase expression of the transcription factor c-fos. In the absence of receptor desensitization, agonist-driven gene expression was suppressed. Mechanistically, stimulation of non-desensitizing receptors evoked prolonged Ca2+ release, which led to accelerated Ca2+-dependent inactivation of CRAC channels and a subsequent loss of excitation-transcription coupling. Rather than serving to turn off a biological response, the experiments show that reversible receptor desensitization is an ‘on’switch’, sustaining long-term signalling in the immune system.EThOS - Electronic Theses Online ServiceGBUnited Kingdo