Structural Basis for the Antiviral Activity of BST-2/Tetherin and Its Viral Antagonism

The interferon-inducible host restriction factor bone marrow stromal antigen 2 (BST-2/tetherin) blocks the release of HIV-1 and other enveloped viruses. In turn, these viruses have evolved specific antagonists to counteract this host antiviral molecule, such as the HIV-1 protein Vpu. BST-2 is a type II transmembrane protein with an unusual topology consisting of an N-terminal cytoplasmic tail (CT) followed by a single transmembrane (TM) domain, a coiled-coil extracellular (EC) domain, and a glycosylphosphatidylinositol (GPI) anchor at the C terminus. We and others showed that BST-2 restricts enveloped virus release by bridging the host and virion membranes with its two opposing membrane anchors and that deletion of either one completely abrogates antiviral activity. The EC domain also shows conserved structural properties that are required for antiviral function. It contains several destabilizing amino acids that confer the molecule with conformational flexibility to sustain the protein’s function as a virion tether, and three conserved cysteine residues that mediate homodimerization of BST-2, as well as acting as a molecular ruler that separates the membrane anchors. Conversely, the efficient release of virions is promoted by the HIV-1 Vpu protein and other viral antagonists. Our group and others provided evidence from mutational analyses indicating that Vpu antagonism of BST-2-mediated viral restriction requires a highly specific interaction of their mutual TM domains. This interpretation is further supported and expanded by the findings of the latest structural modeling studies showing that critical amino acids in a conserved helical face of these TM domains are required for Vpu–BST-2 interaction and antagonism. In this review, we summarize the current advances in our understanding of the structural basis for BST-2 antiviral function as well as BST-2-specific viral antagonism

HIV-1 Vpu Blocks Recycling and Biosynthetic Transport of the Intrinsic Immunity Factor CD317/Tetherin To Overcome the Virion Release Restriction

The intrinsic immunity factor CD317 (BST-2/HM1.24/tetherin) imposes a barrier to HIV-1 release at the cell surface that can be overcome by the viral protein Vpu. Expression of Vpu results in a reduction of CD317 surface levels; however, the mechanism of this Vpu activity and its contribution to the virological antagonism are incompletely understood. Here, we characterized the influence of Vpu on major CD317 trafficking pathways using quantitative antibody-based endocytosis and recycling assays as well as a microinjection/microscopy-based kinetic de novo expression approach. We report that HIV-1 Vpu inhibited both the anterograde transport of newly synthesized CD317 and the recycling of CD317 to the cell surface, while the kinetics of CD317 endocytosis remained unaffected. Vpu trapped trafficking CD317 molecules at the trans-Golgi network, where the two molecules colocalized. The subversion of both CD317 transport pathways was dependent on the highly conserved diserine S52/S56 motif of Vpu; however, it did not require recruitment of the diserine motif interactor and substrate adaptor of the SCF-E3 ubiquitin ligase complex, β-TrCP. Treatment of cells with the malaria drug primaquine resulted in a CD317 trafficking defect that mirrored that induced by Vpu. Importantly, primaquine could functionally replace Vpu as a CD317 antagonist and rescue HIV-1 particle release

Gaps to bridge: Misalignment between perception, reality and actions in obesity

Aims Despite increased recognition as a chronic disease, obesity remains greatly underdiagnosed and undertreated. We aimed to identify international perceptions, attitudes, behaviours and barriers to effective obesity care in people with obesity (PwO) and healthcare professionals (HCPs). Materials and methods An online survey was conducted in 11 countries. Participants were adults with obesity and HCPs who were primarily concerned with direct patient care. Results A total of 14 502 PwO and 2785 HCPs completed the survey. Most PwO (68%) and HCPs (88%) agreed that obesity is a disease. However, 81% of PwO assumed complete responsibility for their own weight loss and only 44% of HCPs agreed that genetics were a barrier. There was a median of three (mean, six) years between the time PwO began struggling with excess weight or obesity and when they first discussed their weight with an HCP. Many PwO were concerned about the impact of excess weight on health (46%) and were motivated to lose weight (48%). Most PwO (68%) would like their HCP to initiate a conversation about weight and only 3% were offended by such a conversation. Among HCPs, belief that patients have little interest in or motivation for weight management may constitute a barrier for weight management conversations. When discussed, HCPs typically recommended lifestyle changes; however, more referrals and follow‐up appointments are required. Conclusions Our international dataset reveals a need to increase understanding of obesity and improve education concerning its physiological basis and clinical management. Realization that PwO are motivated to lose weight offers an opportunity for HCPs to initiate earlier weight management conversations

Sex differences in the regulation of porcine coronary artery tone by perivascular adipose tissue: a role of adiponectin?

Background and Purpose- As there is sexual dimorphism in the regulation of vascular tone, the aim of this present study was to determine whether there are sex differences in perivascular adipose tissue (PVAT) - mediated regulation of the porcine coronary artery (PCA) tone. Experimental Approach- Isometric tension recording system was used to record changes in tone in PCAs. Western blot analysis was performed to examine the expression of adiponectin in PVAT and adiponectin receptors (adipo 1 receptor and adipo 2 receptor) and adiponectin binding protein (APPL1) in PCA. The level of adiponectin released from PVAT was measured using ELISA. Key Results- In the presence of adherent PVAT, contractions to the thromboxane mimetic U46619 and endothelin-1 were significantly reduced in PCAs from females, but not males. In PCAs pre-contracted with U46619, re-addition of PVAT caused relaxation in PCAs from females, but not males. This relaxant response in females was attenuated by combined inhibition of NO synthase (with L-NAME) and cyclooxygenase (with indomethacin). Pre-incubation with an anti-adiponectin antibody abolished the relaxant effects of PVAT. The adiponectin receptor agonist (adipoRon) produced a greater relaxation in PCAs from females compared to males. However, there was no difference in either expression or release of adiponectin from PVAT between sexes. Similarly, there was no difference in expression of adiponectin receptors or the adiponectin receptor adaptor protein APPL1 in PCAs. Conclusion and Implications- These findings demonstrate a clear sex difference in the regulation of coronary artery tone in response to adiponectin receptor stimulation, which may underlie the anticontractile effects of PVAT in females

Coinfection rates in Φ6 bacteriophage are enhanced by virus-induced changes in host cells

Two or more viruses infecting the same host cell can interact in ways that profoundly affect disease dynamics and control, yet the factors determining coinfection rates are incompletely understood. Previous studies have focused on the mechanisms that viruses use to suppress coinfection, but recently the phenomenon of enhanced coinfection has also been documented. In the experiments described here, we explore the hypothesis that enhanced coinfection rates in the bacteriophage Φ6 are achieved by virus-induced upregulation of the Φ6 receptor, which is the bacterial pilus. First, we confirmed that coinfection enhancement in Φ6 is virus-mediated by showing that Φ6 attaches significantly faster to infected cells than to uninfected cells. Second, we explored the hypothesis that coinfection enhancement in Φ6 depends upon changes in the expression of an inducible receptor. Consistent with this hypothesis, the closely related phage, Φ12, that uses constitutively expressed lipopolysaccharide as its receptor, attaches to infected and uninfected cells at the same rate. Our results, along with the previous finding that coinfection in Φ6 is limited to two virions, suggest that viruses may closely regulate rates of coinfection through mechanisms for both coinfection enhancement and exclusion

α-Actinin-4 Is Essential for Maintaining the Spreading, Motility and Contractility of Fibroblasts

Background: α-actinins cross-link actin filaments, with this cross-linking activity regulating the formation of focal adhesions, intracellular tension, and cell migration. Most non-muscle cells such as fibroblasts express two isoforms, α-actinin-1 (ACTN1) and α-actinin-4 (ACTN4). The high homology between these two isoforms would suggest redundancy of their function, but recent studies have suggested different regulatory roles. Interestingly, ACTN4 is phosphorylated upon growth factor stimulation, and this loosens its interaction with actin. Methodology/Principal Findings: Using molecular, biochemical and cellular techniques, we probed the cellular functions of ACTN4 in fibroblasts. Knockdown of ACTN4 expression in murine lung fibroblasts significantly impaired cell migration, spreading, adhesion, and proliferation. Surprisingly, knockdown of ACTN4 enhanced cellular compaction and contraction force, and increased cellular and nuclear cross-sectional area. These results, except the increased contractility, are consistent with a putative role of ACTN4 in cytokinesis. For the transcellular tension, knockdown of ACTN4 significantly increased the expression of myosin light chain 2, a element of the contractility machinery. Re-expression of wild type human ACTN4 in ACTN4 knockdown murine lung fibroblasts reverted cell spreading, cellular and nuclear cross-sectional area, and contractility back towards baseline, demonstrating that the defect was due to absence of ACTN4. Significance: These results suggest that ACTN4 is essential for maintaining normal spreading, motility, cellular and nuclear cross-sectional area, and contractility of murine lung fibroblasts by maintaining the balance between transcellular contractility and cell-substratum adhesion. © 2010 Shao et al

PKCδ regulates force signaling during VEGF/CXCL4 induced dissociation of endothelial tubes

Wound healing requires the vasculature to re-establish itself from the severed ends; endothelial cells within capillaries must detach from neighboring cells before they can migrate into the nascent wound bed to initiate angiogenesis. The dissociation of these endothelial capillaries is driven partially by platelets' release of growth factors and cytokines, particularly the chemokine CXCL4/platelet factor-4 (PF4) that increases cell-cell de-adherence. As this retraction is partly mediated by increased transcellular contractility, the protein kinase c-δ/myosin light chain-2 (PKCδ/MLC-2) signaling axis becomes a candidate mechanism to drive endothelial dissociation. We hypothesize that PKCδ activation induces contractility through MLC-2 to promote dissociation of endothelial cords after exposure to platelet-released CXCL4 and VEGF. To investigate this mechanism of contractility, endothelial cells were allowed to form cords following CXCL4 addition to perpetuate cord dissociation. In this study, CXCL4-induced dissociation was reduced by a VEGFR inhibitor (sunitinib malate) and/or PKCδ inhibition. During combined CXCL4+VEGF treatment, increased contractility mediated by MLC-2 that is dependent on PKCδ regulation. As cellular force is transmitted to focal adhesions, zyxin, a focal adhesion protein that is mechano-responsive, was upregulated after PKCδ inhibition. This study suggests that growth factor regulation of PKCδ may be involved in CXCL4-mediated dissociation of endothelial cords. © 2014 Jamison et al

The role of adiponectin receptors in the regulation of synaptic transmission in the hippocampus

In the last two decades adiponectin, member of the adipokines family, gained attention because of its unique antidiabetic effects. However, the presence in the brain of adiponectin receptors and adiponectin itself raised interest because of the possible association with neuropsychiatric diseases. Indeed, clinical studies found altered concentration of adiponectin both in plasma and cerebrospinal fluid in seeveral pathologies including depression, multiple sclerosis, Alzheimer’s disease and stroke. Moreover, recent preclinical studies also suggest its involvement in different physiological functions. Despite this evidence very few studies attempted to elucidate the functional role of adiponectin at the synapse.To address this question, here we investigated the effect of Adiporon, an agonist of both adiponectin receptors on synaptic transmission and LTP at Schaffer-collateral CA1 pathway. Surprisingly, increasing concentration of Adiporon correlated with lower CA1-LTP levels and paired-pulse ratio, whereas basal transmission was always preserved. Collectively, our data show that the adiponectin system, beyond its involvement in metabolic diseases, plays also a critical role in synaptic activity thereby representing a putative target for the treatment of synaptic pathologies