Phospholipids: pulling back the actin curtain for granule delivery to the immune synapse

Phosphoinositides, together with the phospholipids phosphatidylserine and phosphatidic acid, are important components of the plasma membrane acting as second messengers that, with diacylglycerol, regulate a diverse range of signalling events converting extracellular changes into cellular responses. Local changes in their distribution and membrane charge on the inner leaflet of the plasma membrane play important roles in immune cell function. Here we discuss their distribution and regulators highlighting the importance of membrane changes across the immune synapse on the cytoskeleton and the impact on the function of cytotoxic T lymphocytes.Research was funded by Wellcome Trust grants [103930] and [100140], to GMG

Phospholipids: Pulling Back the Actin Curtain for Granule Delivery to the Immune Synapse

Phosphoinositides, together with the phospholipids phosphatidylserine and phosphatidic acid, are important components of the plasma membrane acting as second messengers that, with diacylglycerol, regulate a diverse range of signaling events converting extracellular changes into cellular responses. Local changes in their distribution and membrane charge on the inner leaflet of the plasma membrane play important roles in immune cell function. Here we discuss their distribution and regulators highlighting the importance of membrane changes across the immune synapse on the cytoskeleton and the impact on the function of cytotoxic T lymphocytes

Loss of ARPC1B impairs cytotoxic T lymphocyte maintenance and cytolytic activity.

CD8 cytotoxic T lymphocytes (CTLs) rely on rapid reorganization of the branched F-actin network to drive the polarized secretion of lytic granules, initiating target cell death during the adaptive immune response. Branched F-actin is generated by the nucleation factor actin-related protein 2/3 (Arp2/3) complex. Patients with mutations in the actin-related protein complex 1B (ARPC1B) subunit of Arp2/3 show combined immunodeficiency, with symptoms of immune dysregulation, including recurrent viral infections and reduced CD8+ T cell count. Here, we show that loss of ARPC1B led to loss of CTL cytotoxicity, with the defect arising at 2 different levels. First, ARPC1B is required for lamellipodia formation, cell migration, and actin reorganization across the immune synapse. Second, we found that ARPC1B is indispensable for the maintenance of TCR, CD8, and GLUT1 membrane proteins at the plasma membrane of CTLs, as recycling via the retromer and WASH complexes was impaired in the absence of ARPC1B. Loss of TCR, CD8, and GLUT1 gave rise to defects in T cell signaling and proliferation upon antigen stimulation of ARPC1B-deficient CTLs, leading to a progressive loss of CD8+ T cells. This triggered an activation-induced immunodeficiency of CTL activity in ARPC1B-deficient patients, which could explain the susceptibility to severe and prolonged viral infections

Podosomes of dendritic cells facilitate antigen sampling

Dendritic cells sample the environment for antigens and play an important role in establishing the link between innate and acquired immunity. Dendritic cells contain mechanosensitive adhesive structures called podosomes that consist of an actin-rich core surrounded by integrins, adaptor proteins and actin network filaments. They facilitate cell migration via localized degradation of extracellular matrix. Here, we show that podosomes of human dendritic cells locate to spots of low physical resistance in the substrate (soft spots) where they can evolve into protrusive structures. Pathogen recognition receptors locate to these protrusive structures where they can trigger localized antigen uptake, processing and presentation to activate T-cells. Our data demonstrate a novel role in antigen sampling for the podosomes of dendritic cell

Actin depletion initiates events leading to granule secretion at the immunological synapse.

Cytotoxic T lymphocytes (CTLs) use polarized secretion to rapidly destroy virally infected and tumor cells. To understand the temporal relationships between key events leading to secretion, we used high-resolution 4D imaging. CTLs approached targets with actin-rich projections at the leading edge, creating an initially actin-enriched contact with rearward-flowing actin. Within 1 min, cortical actin reduced across the synapse, T cell receptors (TCRs) clustered centrally to form the central supramolecular activation cluster (cSMAC), and centrosome polarization began. Granules clustered around the moving centrosome within 2.5 min and reached the synapse after 6 min. TCR-bearing intracellular vesicles were delivered to the cSMAC as the centrosome docked. We found that the centrosome and granules were delivered to an area of membrane with reduced cortical actin density and phospholipid PIP2. These data resolve the temporal order of events during synapse maturation in 4D and reveal a critical role for actin depletion in regulating secretion.Funding was provided by the Wellcome Trust through Principal Research Fellowships (075880 and 103930) to G.M.G. and a Strategic Award (100140) to the Cambridge Institute for Medical Research (CIMR). A.T.R. is an NIH-OxCam scholar supported by funding to J.L.-S. from the Eunice Shriver National Institute of Child Health and Human Development.This is the final version. It was first published by Elsevier at http://www.cell.com/immunity/abstract/S1074-7613%2815%2900173-9

Multicentric osteolytic syndromes represent a phenotypic spectrum defined by defective collagen remodeling

Frank-Ter Haar syndrome (FTHS), Winchester syndrome (WS), and multicentric osteolysis, nodulosis, and arthropathy (MONA) are ultra-rare multisystem disorders characterized by craniofacial malformations, reduced bone density, skeletal and cardiac anomalies, and dermal fibrosis. These autosomal recessive syndromes are caused by homozygous mutation or deletion of respectively SH3PXD2B (SH3 and PX Domains 2B), MMP14 (matrix metalloproteinase 14), or MMP2. Here, we give an overview of the clinical features of 63 previously reported patients with an SH3PXD2B, MMP14, or MMP2 mutation, demonstrating considerable clinical overlap between FTHS, WS, and MONA. Interestingly, the protein products of SH3PXD2B, MMP14, and MMP2 directly cooperate in collagen remodeling. We review animal models for these three disorders that accurately reflect the major clinical features and likewise show significant phenotypical similarity with each other. Furthermore, they demonstrate that defective collagen remodeling is central in the underlying pathology. As such, we propose a nosological revision, placing these SH3PXD2B, MMP14, and MMP2 related syndromes in a novel “defective collagen-remodelling spectrum (DECORS)”. In our opinion, this revised nosology better reflects the central role for impaired collagen remodeling, a potential target for pharmaceutical intervention

Geometry sensing by dendritic cells dictates spatial organization and PGE2-induced dissolution of podosomes

Assembly and disassembly of adhesion structures such as focal adhesions (FAs) and podosomes regulate cell adhesion and differentiation. On antigen-presenting dendritic cells (DCs), acquisition of a migratory and immunostimulatory phenotype depends on podosome dissolution by prostaglandin E2 (PGE2). Whereas the effects of physico-chemical and topographical cues have been extensively studied on FAs, little is known about how podosomes respond to these signals. Here, we show that, unlike for FAs, podosome formation is not controlled by substrate physico-chemical properties. We demonstrate that cell adhesion is the only prerequisite for podosome formation and that substrate availability dictates podosome density. Interestingly, we show that DCs sense 3-dimensional (3-D) geometry by aligning podosomes along the edges of 3-D micropatterned surfaces. Finally, whereas on a 2-dimensional (2-D) surface PGE2 causes a rapid increase in activated RhoA levels leading to fast podosome dissolution, 3-D geometric cues prevent PGE2-mediated RhoA activation resulting in impaired podosome dissolution even after prolonged stimulation. Our findings indicate that 2-D and 3-D geometric cues control the spatial organization of podosomes. More importantly, our studies demonstrate the importance of substrate dimensionality in regulating podosome dissolution and suggest that substrate dimensionality plays an important role in controlling DC activation, a key process in initiating immune responses

Ultrastructural characterisation of the dendritic cell podosome as a means to understand podosome function

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