Exercise hormone irisin is a critical regulator of cognitive function.

Identifying secreted mediators that drive the cognitive benefits of exercise holds great promise for the treatment of cognitive decline in ageing or Alzheimer's disease (AD). Here, we show that irisin, the cleaved and circulating form of the exercise-induced membrane protein FNDC5, is sufficient to confer the benefits of exercise on cognitive function. Genetic deletion of Fndc5/irisin (global Fndc5 knock-out (KO) mice; F5KO) impairs cognitive function in exercise, ageing and AD. Diminished pattern separation in F5KO mice can be rescued by delivering irisin directly into the dentate gyrus, suggesting that irisin is the active moiety. In F5KO mice, adult-born neurons in the dentate gyrus are morphologically, transcriptionally and functionally abnormal. Importantly, elevation of circulating irisin levels by peripheral delivery of irisin via adeno-associated viral overexpression in the liver results in enrichment of central irisin and is sufficient to improve both the cognitive deficit and neuropathology in AD mouse models. Irisin is a crucial regulator of the cognitive benefits of exercise and is a potential therapeutic agent for treating cognitive disorders including AD.

Thioredoxin Interacting Protein Is Required for a Chronic Energy-Rich Diet to Promote Intestinal Fructose Absorption.

Increased consumption of fats and added sugars has been associated with an increase in metabolic syndromes. Here we show that mice chronically fed an energy-rich diet (ERD) with high fat and moderate sucrose have enhanced the absorption of a gastrointestinal fructose load, and this required expression of the arrestin domain protein Txnip in the intestinal epithelial cells. ERD feeding induced gene and protein expression of Glut5, and this required the expression of Txnip. Furthermore, Txnip interacted with Rab11a, a small GTPase that facilitates the apical localization of Glut5. We also demonstrate that ERD promoted Txnip/Glut5 complexes in the apical intestinal epithelial cell. Our findings demonstrate that ERD facilitates fructose absorption through a Txnip-dependent mechanism in the intestinal epithelial cell, suggesting that increased fructose absorption could potentially provide a mechanism for worsening of metabolic syndromes in the setting of a chronic ERD.

Two- and three-color STORM analysis reveals higher-order assembly of leukotriene synthetic complexes on the nuclear envelope of murine neutrophils.

Over the last several years it has become clear that higher order assemblies on membranes, exemplified by signalosomes, are a paradigm for the regulation of many membrane signaling processes. We have recently combined two-color direct stochastic optical reconstruction microscopy (dSTORM) with the (Clus-DoC) algorithm that combines cluster detection and colocalization analysis to observe the organization of 5-lipoxygenase (5-LO) and 5-lipoxygenase-activating protein (FLAP) into higher order assemblies on the nuclear envelope of mast cells; these assemblies were linked to leukotriene (LT) C4 production. In this study we investigated whether higher order assemblies of 5-LO and FLAP included cytosolic phospholipase A2 (cPLA2) and were linked to LTB4 production in murine neutrophils. Using two- and three-color dSTORM supported by fluorescence lifetime imaging microscopy we identified higher order assemblies containing 40 molecules (median) (IQR: 23, 87) of 5-LO, and 53 molecules (62, 156) of FLAP monomer. 98 (18, 154) molecules of cPLA2 were clustered with 5-LO, and 77 (33, 114) molecules of cPLA2 were associated with FLAP. These assemblies were tightly linked to LTB4 formation. The activation-dependent close associations of cPLA2, FLAP, and 5-LO in higher order assemblies on the nuclear envelope support a model in which arachidonic acid is generated by cPLA2 in apposition to FLAP, facilitating its transfer to 5-LO to initiate LT synthesis.

The organization of leukotriene biosynthesis on the nuclear envelope revealed by single molecule localization microscopy and computational analyses.

The initial steps in the synthesis of leukotrienes are the translocation of 5-lipoxygenase (5-LO) to the nuclear envelope and its subsequent association with its scaffold protein 5-lipoxygenase-activating protein (FLAP). A major gap in our understanding of this process is the knowledge of how the organization of 5-LO and FLAP on the nuclear envelope regulates leukotriene synthesis. We combined single molecule localization microscopy with Clus-DoC cluster analysis, and also a novel unbiased cluster analysis to analyze changes in the relationships between 5-LO and FLAP in response to activation of RBL-2H3 cells to generate leukotriene C4. We identified the time-dependent reorganization of both 5-LO and FLAP into higher-order assemblies or clusters in response to cell activation via the IgE receptor. Clus-DoC analysis identified a subset of these clusters with a high degree of interaction between 5-LO and FLAP that specifically correlates with the time course of LTC4 synthesis, strongly suggesting their role in the initiation of leukotriene biosynthesis.

Differential attenuation of ß2 integrin-dependent and -independent neutrophil migration by Ly6G ligation.

Antibody ligation of the murine neutrophil surface protein Ly6G disrupts neutrophil migration in some contexts but not others. We tested whether this variability reflected divergent dependence of neutrophil migration on ß2 integrins, adhesion molecules that interact with Ly6G at the neutrophil surface. In integrin-dependent murine arthritis, Ly6G ligation attenuated joint inflammation, even though mice lacking Ly6G altogether developed arthritis normally. By contrast, Ly6G ligation had no impact on integrin-independent neutrophil migration into inflamed lung. In peritoneum, the role of ß2 integrins varied with stimulus, proving dispensable for neutrophil entry in Escherichia coli peritonitis but contributory in interleukin 1 (IL-1)-mediated sterile peritonitis. Correspondingly, Ly6G ligation attenuated only IL-1 peritonitis, disrupting the molecular association between integrins and Ly6G and inducing cell-intrinsic blockade restricted to integrin-dependent migration. Consistent with this observation, Ly6G ligation impaired integrin-mediated postadhesion strengthening for neutrophils arresting on activated cremaster endothelium in vivo. Together, these findings identify selective inhibition of integrin-mediated neutrophil emigration through Ly6G ligation, highlighting the marked site and stimulus specificity of ß2 integrin dependence in neutrophil migration.

CD177 modulates human neutrophil migration through activation-mediated integrin and chemoreceptor regulation.

CD177 is a glycosylphosphatidylinositol (GPI)-anchored protein expressed by a variable proportion of human neutrophils that mediates surface expression of the antineutrophil cytoplasmic antibody antigen proteinase 3. CD177 associates with ß2 integrins and recognizes platelet endothelial cell adhesion molecule 1 (PECAM-1), suggesting a role in neutrophil migration. However, CD177pos neutrophils exhibit no clear migratory advantage in vivo, despite interruption of in vitro transendothelial migration by CD177 ligation. We sought to understand this paradox. Using a PECAM-1-independent transwell system, we found that CD177pos and CD177neg neutrophils migrated comparably. CD177 ligation selectively impaired migration of CD177pos neutrophils, an effect mediated through immobilization and cellular spreading on the transwell membrane. Correspondingly, CD177 ligation enhanced its interaction with ß2 integrins, as revealed by fluorescence lifetime imaging microscopy, leading to integrin-mediated phosphorylation of Src and extracellular signal-regulated kinase (ERK). CD177-driven cell activation enhanced surface ß2 integrin expression and affinity, impaired internalization of integrin attachments, and resulted in ERK-mediated attenuation of chemokine signaling. We conclude that CD177 signals in a ß2 integrin-dependent manner to orchestrate a set of activation-mediated mechanisms that impair human neutrophil migration.

Diabetes regulates fructose absorption through thioredoxin-interacting protein.

Metabolic studies suggest that the absorptive capacity of the small intestine for fructose is limited, though the molecular mechanisms controlling this process remain unknown. Here we demonstrate that thioredoxin-interacting protein (Txnip), which regulates glucose homeostasis in mammals, binds to fructose transporters and promotes fructose absorption by the small intestine. Deletion of Txnip in mice reduced fructose transport into the peripheral bloodstream and liver, as well as the severity of adverse metabolic outcomes resulting from long-term fructose consumption. We also demonstrate that fructose consumption induces expression of Txnip in the small intestine. Diabetic mice had increased expression of Txnip in the small intestine as well as enhanced fructose uptake and transport into the hepatic portal circulation. The deletion of Txnip in mice abolished the diabetes-induced increase in fructose absorption. Our results indicate that Txnip is a critical regulator of fructose metabolism and suggest that a diabetic state can promote fructose uptake.

Coordinated regulation of bidirectional COPI transport at the Golgi by CDC42.

The Golgi complex has a central role in the intracellular sorting of secretory proteins. Anterograde transport through the Golgi has been explained by the movement of Golgi cisternae, known as cisternal maturation. Because this explanation is now appreciated to be incomplete, interest has developed in understanding tubules that connect the Golgi cisternae. Here we show that the coat protein I (COPI) complex sorts anterograde cargoes into these tubules in human cells. Moreover, the small GTPase CDC42 regulates bidirectional Golgi transport by targeting the dual functions of COPI in cargo sorting and carrier formation. CDC42 also directly imparts membrane curvature to promote COPI tubule formation. Our findings further reveal that COPI tubular transport complements cisternal maturation in explaining how anterograde Golgi transport is achieved, and that bidirectional COPI transport is modulated by environmental cues through CDC42.

CD47 plays a critical role in T-cell recruitment by regulation of LFA-1 and VLA-4 integrin adhesive functions.

CD47 plays an important but incompletely understood role in the innate and adaptive immune responses. CD47, also called integrin-associated protein, has been demonstrated to associate in cis with ß1 and ß3 integrins. Here we test the hypothesis that CD47 regulates adhesive functions of T-cell α4ß1 (VLA-4) and αLß2 (LFA-1) in in vivo and in vitro models of inflammation. Intravital microscopy studies reveal that CD47(-/-) Th1 cells exhibit reduced interactions with wild-type (WT) inflamed cremaster muscle microvessels. Similarly, murine CD47(-/-) Th1 cells, as compared with WT, showed defects in adhesion and transmigration across tumor necrosis factor-α (TNF-α)-activated murine endothelium and in adhesion to immobilized intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion protein 1 (VCAM-1) under flow conditions. Human Jurkat T-cells lacking CD47 also showed reduced adhesion to TNF-α-activated endothelium and ICAM-1 and VCAM-1. In cis interactions between Jurkat T-cell ß2 integrins and CD47 were detected by fluorescence lifetime imaging microscopy. Unexpectedly, Jurkat CD47 null cells exhibited a striking defect in ß1 and ß2 integrin activation in response to Mn(2+) or Mg(2+)/ethylene glycol tetraacetic acid treatment. Our results demonstrate that CD47 associates with ß2 integrins and is necessary to induce high-affinity conformations of LFA-1 and VLA-4 that recognize their endothelial cell ligands and support leukocyte adhesion and transendothelial migration.