Phosphoinositide lipids and cell polarity: linking the plasma membrane to the cytocortex

2012 ◽  
Vol 53 ◽  
pp. 15-27 ◽  
Author(s):  
Michael P. Krahn ◽  
Andreas Wodarz
Keyword(s):  
Plasma Membrane ◽  
Epithelial Cells ◽  
Membrane Lipids ◽  
Leading Edge ◽  
Cell Types ◽  
Specific Protein ◽  
Molecular Organization ◽  
Apical Side ◽  
Cell Substrate ◽  
Basolateral Side

Many cell types in animals and plants are polarized, which means that the cell is subdivided into functionally and structurally distinct compartments. Epithelial cells, for example, possess an apical side facing a lumen or the outside environment and a basolateral side facing adjacent epithelial cells and the basement membrane. Neurons possess distinct axonal and dendritic compartments with specific functions in sending and receiving signals. Migrating cells form a leading edge that actively engages in pathfinding and cell-substrate attachment, and a trailing edge where such attachments are abandoned. In all of these cases, both the plasma membrane and the cytocortex directly underneath the plasma membrane show differences in their molecular composition and structural organization. In this chapter we will focus on a specific type of membrane lipids, the phosphoinositides, because in polarized cells they show a polarized distribution in the plasma membrane. They furthermore influence the molecular organization of the cytocortex by recruiting specific protein binding partners which are involved in the regulation of the cytoskeleton and in signal transduction cascades that control polarity, growth and cell migration.

scholarly journals ALIX and ceramide differentially control polarized exosome release from epithelial cells

2020 ◽  
Author(s):  
Takahide Matsui ◽  
Shu Hiragi ◽  
Futaba Osaki ◽  
Yuriko Sakamaki ◽  
Mitsunori Fukuda
Keyword(s):  
Epithelial Cells ◽  
Single Cells ◽  
Cell Communication ◽  
Specific Protein ◽  
Endosomal Sorting ◽  
Apical Side ◽  
Basolateral Side ◽  
Group 5 ◽  
G Protein Coupled ◽  
Ceramide Production

AbstractExosomes, new players in cell-cell communication, are extracellular vesicles of endocytic origin. Although single cells are known to release various kinds of exosomes (referred to as exosomal heterogeneity), very little is known about the mechanisms by which they are produced and released. Here, we established methods for studying exosomal heterogeneity by using polarized epithelial cells and showed that distinct types of exosomes are differentially secreted from the apical and basolateral sides. We also identified GPRC5C (G protein-coupled receptor class C group 5 member C) as an apical-exosome-specific protein. We further demonstrated that basolateral exosome release depends on ceramide, whereas ALIX, an ESCRT (endosomal sorting complexes required for transport)-related protein, not the ESCRT machinery itself, is required for apical exosome secretion. Thus, two independent machineries, the ALIX–Syntenin1– Syndecan1 machinery (apical side) and the sphingomyelinase-dependent ceramide production machinery (basolateral side), are likely to be responsible for the polarized exosome release from epithelial cells.

Membrane microdomains: lessons from microscopy

1995 ◽  
Vol 53 ◽  
pp. 776-777
Author(s):  
J.M. Robinson ◽  
J.M Oliver
Keyword(s):  
Spatial Distribution ◽  
Plasma Membrane ◽  
Epithelial Cells ◽  
Plasma Membranes ◽  
Cell Types ◽  
Imaging Modalities ◽  
Membrane Microdomains ◽  
Membrane Domains ◽  
Lateral Heterogeneity ◽  
Functional Importance

Specialized regions of plasma membranes displaying lateral heterogeneity are the focus of this Symposium. Specialized membrane domains are known for certain cell types such as differentiated epithelial cells where lateral heterogeneity in lipids and proteins exists between the apical and basolateral portions of the plasma membrane. Lateral heterogeneity and the presence of microdomains in membranes that are uniform in appearance have been more difficult to establish. Nonetheless a number of studies have provided evidence for membrane microdomains and indicated a functional importance for these structures.This symposium will focus on the use of various imaging modalities and related approaches to define membrane microdomains in a number of cell types. The importance of existing as well as emerging imaging technologies for use in the elucidation of membrane microdomains will be highlighted. The organization of membrane microdomains in terms of dimensions and spatial distribution is of considerable interest and will be addressed in this Symposium.

Nanotopography-Modulated Epithelial Cell Collective Migration

2021 ◽  
Vol 17 (6) ◽  
pp. 1079-1087
Author(s):  
Zaozao Chen ◽  
Qiwei Li ◽  
Shihui Xu ◽  
Jun Ouyang ◽  
Hongmei Wei
Keyword(s):  
Wound Healing ◽  
Cell Migration ◽  
Epithelial Cells ◽  
Leading Edge ◽  
Cell Types ◽  
Protein Kinase Activity ◽  
Migration Behavior ◽  
Collective Migration ◽  
Epithelial Migration ◽  
Activity Dependent

Matrix nanotopography plays an essential role in regulating cell behaviors including cell proliferation, differentiation, and migration. While studies on isolated single cell migration along the nanostructural orientation have been reported for various cell types, there remains a lack of understanding of how nanotopography regulates the behavior of collectively migrating cells during processes such as epithelial wound healing. We demonstrated that collective migration of epithelial cells was promoted on nanogratings perpendicular to, but not on those parallel to, the wound-healing axis. We further discovered that nanograting-modulated epithelial migration was dominated by the adhesion turnover process, which was Rho-associated protein kinase activity-dependent, and the lamellipodia protrusion at the cell leading edge, which was Rac1-GTPase activity-dependent. This work provides explanations to the distinct migration behavior of epithelial cells on nanogratings, and indicates that the effect of nanotopographic modulations on cell migration is cell-type dependent and involves complex mechanisms

scholarly journals A Note of Caution: Gramicidin Affects Signaling Pathways Independently of Its Effects on Plasma Membrane Conductance

2021 ◽  
Vol 2021 ◽  
pp. 1-12
Author(s):  
Frances Evans ◽  
Julio A. Hernández ◽  
Federico Cabo ◽  
Silvia Chifflet
Keyword(s):  
Plasma Membrane ◽  
Membrane Potential ◽  
Electrical Properties ◽  
Epithelial Cells ◽  
Molecular Mechanisms ◽  
Membrane Lipids ◽  
Membrane Conductance ◽  
Ionic Channel ◽  
Plasma Membrane Potential ◽  
Hydrophobic Nature

Gramicidin is a thoroughly studied cation ionophore widely used to experimentally manipulate the plasma membrane potential (PMP). In addition, it has been established that the drug, due to its hydrophobic nature, is capable of affecting the organization of membrane lipids. We have previously shown that modifications in the plasma membrane potential of epithelial cells in culture determine reorganizations of the cytoskeleton. To elucidate the molecular mechanisms involved, we explored the effects of PMP depolarization on some putative signaling intermediates. In the course of these studies, we came across some results that could not be interpreted in terms of the properties of gramicidin as an ionic channel. The purpose of the present work is to communicate these results and, in general, to draw attention to the fact that gramicidin effects can be misleadingly attributed to its ionic or electrical properties. In addition, this work also contributes with some novel findings of the modifications provoked on the signaling intermediates by PMP depolarization and hyperpolarization.

scholarly journals Polarity sensitive probes for super resolution STED microscopy

2017 ◽  
Author(s):  
E Sezgin ◽  
F Schneider ◽  
V Zilles ◽  
E Garcia ◽  
D Waithe ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Lipid Membrane ◽  
Membrane Lipids ◽  
Super Resolution ◽  
Membrane Vesicles ◽  
Molecular Organization ◽  
Live Cells ◽  
Sted Microscopy ◽  
Two Photon Microscopy ◽  
Polarity Sensitive

AbstractThe lateral organization of molecules in the cellular plasma membrane plays an important role in cellular signaling. A critical parameter for membrane molecular organization is how the membrane lipids are packed (or ordered). Polarity sensitive dyes are powerful tools to characterize such lipid membrane order, employing for example confocal and two-photon microscopy. The investigation of potential lipid nanodomains, however, requires the use of super resolution microscopy. Here, we test the performance of the polarity sensitive membrane dyes Di-4-ANEPPDHQ, Di-4-AN(F)EPPTEA and NR12S in super resolution STED microscopy. Measurements on cell-derived membrane vesicles, in the plasma membrane of live cells, and on single virus particles show the high potential of these dyes for probing nanoscale membrane heterogeneity.

scholarly journals Dynamics of a Chemoattractant Receptor in Living Neutrophils during Chemotaxis

1999 ◽  
Vol 10 (4) ◽  
pp. 1163-1178 ◽  
Author(s):  
Guy Servant ◽  
Orion D. Weiner ◽  
Enid R. Neptune ◽  
John W. Sedat ◽  
Henry R. Bourne
Keyword(s):  
Plasma Membrane ◽  
Fluorescent Protein ◽  
Membrane Lipids ◽  
Leukemia Cell ◽  
Leading Edge ◽  
Relative Strength ◽  
Leukemia Cell Line ◽  
Anterior Pole ◽  
Apparent Increase ◽  
Green Fluorescent

Persistent directional movement of neutrophils in shallow chemotactic gradients raises the possibility that cells can increase their sensitivity to the chemotactic signal at the front, relative to the back. Redistribution of chemoattractant receptors to the anterior pole of a polarized neutrophil could impose asymmetric sensitivity by increasing the relative strength of detected signals at the cell’s leading edge. Previous experiments have produced contradictory observations with respect to receptor location in moving neutrophils. To visualize a chemoattractant receptor directly during chemotaxis, we expressed a green fluorescent protein (GFP)-tagged receptor for a complement component, C5a, in a leukemia cell line, PLB-985. Differentiated PLB-985 cells, like neutrophils, adhere, spread, and polarize in response to a uniform concentration of chemoattractant, and orient and crawl toward a micropipette containing chemoattractant. Recorded in living cells, fluorescence of the tagged receptor, C5aR–GFP, shows no apparent increase anywhere on the plasma membrane of polarized and moving cells, even at the leading edge. During chemotaxis, however, some cells do exhibit increased amounts of highly folded plasma membrane at the leading edge, as detected by a fluorescent probe for membrane lipids; this is accompanied by an apparent increase of C5aR–GFP fluorescence, which is directly proportional to the accumulation of plasma membrane. Thus neutrophils do not actively concentrate chemoattractant receptors at the leading edge during chemotaxis, although asymmetrical distribution of membrane may enrich receptor number, relative to adjacent cytoplasmic volume, at the anterior pole of some polarized cells. This enrichment could help to maintain persistent migration in a shallow gradient of chemoattractant.

scholarly journals Examining the Role of Actin-Plasma Membrane Association in Pseudomonas aeruginosa Infection and Type III Secretion Translocation in Migratory T24 Epithelial Cells

2012 ◽  
Vol 80 (9) ◽  
pp. 3049-3064 ◽  
Author(s):  
Dacie R. Bridge ◽  
Karen H. Martin ◽  
Elizabeth R. Moore ◽  
Wendy M. Lee ◽  
James A. Carroll ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Plasma Membrane ◽  
Epithelial Cells ◽  
Type Iii Secretion ◽  
Actin Filaments ◽  
Leading Edge ◽  
Membrane Association ◽  
Wild Type ◽  
Content Type ◽  
T24 Cells

ABSTRACTThe opportunistic pathogenPseudomonas aeruginosatargets wounded epithelial barriers, but the cellular alteration that increases susceptibility toP. aeruginosainfection remains unclear. This study examined how cell migration contributes to the establishment ofP. aeruginosainfections using (i) highly migratory T24 epithelial cells as a cell culture model, (ii) mutations in the type III secretion (T3S) effector ExoS to manipulateP. aeruginosainfection, and (iii) high-resolution immunofluorescent microscopy to monitor ExoS translocation. ExoS includes both GTPase-activating (GAP) and ADP-ribosyltransferase (ADPRT) activities, andP. aeruginosacells expressing wild-type ExoS preferentially bound to the leading edge of T24 cells, where ExoS altered leading-edge architecture and actin anchoring in conjunction with interrupting T3S translocation. Inactivation of ExoS GAP activity allowedP. aeruginosato be internalized and secrete ExoS within T24 cells, but as with wild-type ExoS, translocation was limited in association with disruption of actin anchoring. Inactivation of ExoS ADPRT activity resulted in significantly enhanced T3S translocation byP. aeruginosacells that remained extracellular and in conjunction with maintenance of actin-plasma membrane association. Infection withP. aeruginosaexpressing ExoS lacking both GAP and ADPRT activities resulted in the highest level of T3S translocation, and this occurred in conjunction with the entry and alignment ofP. aeruginosaand ExoS along actin filaments. Collectively, in using ExoS mutants to modulate and visualize T3S translocation, we were able to (i) confirm effector secretion by internalizedP. aeruginosa, (ii) differentiate the mechanisms underlying the effects of ExoS GAP and ADPRT activities onP. aeruginosainternalization and T3S translocation, (iii) confirm that ExoS ADPRT activity targeted a cellular substrate that interrupted T3S translocation, (iv) visualize the ability ofP. aeruginosaand ExoS to align with actin filaments, and (v) demonstrate an association between actin anchoring at the leading edge of T24 cells and the establishment ofP. aeruginosainfection. Our studies also highlight the contribution of ExoS to the opportunistic nature ofP. aeruginosainfection through its ability to exert cytotoxic effects that interrupt T3S translocation andP. aeruginosainternalization, which in turn limit theP. aeruginosainfectious process.

scholarly journals Functional analysis of OCTN2 and ATB0,+ in normal human airway epithelial cells

2019 ◽  
Author(s):  
Bianca Maria Rotoli ◽  
Rossana Visigalli ◽  
Amelia Barilli ◽  
Francesca Ferrari ◽  
Massimiliano G. Bianchi ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Pulmonary Delivery ◽  
Airway Epithelial Cells ◽  
Airway Epithelial ◽  
Apical Side ◽  
Basolateral Side ◽  
Subsequent Degradation ◽  
Carnitine Transport ◽  
Air Liquid Interface

ABSTRACTIn human, OCTN2 (SLC22A5) and ATB0,+ (SLC6A14) transporters mediate the uptake of L-carnitine, essential for the transport of fatty acids into mitochondria and the subsequent degradation by β-oxidation. Aim of the present study is to characterize L-carnitine transport in EpiAirway™, a 3D organotypic in vitro model of primary human tracheal-bronchial epithelial cells that form a fully differentiated, pseudostratified columnar epithelium at air-liquid interface (ALI) condition. In parallel, Calu-3 monolayers grown at ALI were used as comparison. In EpiAirway™, ATB0,+ was highly expressed and functional on the apical side while OCTN2 transporter was active on the basolateral side. Calu-3 cells showed a different pattern of expression and activity for ATB0,+: indeed, L-carnitine uptake on apical side was evident in Calu-3 at 8 days of culture but not in fully differentiated 21d ALI culture. As both ATB0,+ and OCTN2, beyond transporting L-carnitine, have a significant potential as delivery systems for drugs, the identification of these transporters in EpiAirway™ can open new fields of investigation in the studies of drug inhalation and pulmonary delivery.

Structure of the Plasma Membrane of Uterine Epithelial Cells in Blastocyst Attachment: a Review

1992 ◽  
Vol 4 (6) ◽  
pp. 633 ◽  
Author(s):  
CR Murphy
Keyword(s):  
Plasma Membrane ◽  
Epithelial Cells ◽  
Tight Junction ◽  
Freeze Fracture ◽  
Membrane Skeleton ◽  
Cholesterol Content ◽  
Molecular Organization ◽  
Membrane Cholesterol ◽  
Uterine Receptivity ◽  
Uterine Epithelial Cells

Changes in the molecular organization of the plasma membrane of uterine epithelial cells during early pregnancy and, in particular, at the attachment period are reviewed. The review focuses on attachment in rodents but other species are also considered. Alterations in protein content and type, as determined with electrophoretic and freeze-fracture techniques, and an increase in tight junction complexity occur in several species. Ultrastructural histochemistry shows that glycocalyx carbohydrates of different species both increase and decrease depending on the type of carbohydrate. Changes in membrane cholesterol content also occur and recent studies showing major reorganization of the actin-containing membrane skeleton are reviewed to show the dynamism of this plasma membrane during the period of uterine receptivity for attachment of the blastocyst.

scholarly journals Diffusion Coefficient of Fluorescent Phosphatidylinositol 4,5-bisphosphate in the Plasma Membrane of Cells

2008 ◽  
Vol 19 (4) ◽  
pp. 1663-1669 ◽  
Author(s):  
Urszula Golebiewska ◽  
Marian Nyako ◽  
William Woturski ◽  
Irina Zaitseva ◽  
Stuart McLaughlin
Keyword(s):  
Diffusion Coefficient ◽  
Plasma Membrane ◽  
Epithelial Cells ◽  
Fluorescence Correlation Spectroscopy ◽  
Plasma Membranes ◽  
Diffusion Constant ◽  
Cell Types ◽  
Correlation Spectroscopy ◽  
Fluorescence Correlation ◽  
Outer Leaflet

Phosphatidylinositol 4,5-bisphosphate (PIP2) controls a surprisingly large number of processes in cells. Thus, many investigators have suggested that there might be different pools of PIP2 on the inner leaflet of the plasma membrane. If a significant fraction of PIP2 is bound electrostatically to unstructured clusters of basic residues on membrane proteins, the PIP2 diffusion constant, D, should be reduced. We microinjected micelles of Bodipy TMR-PIP2 into cells, and we measured D on the inner leaflet of fibroblasts and epithelial cells by using fluorescence correlation spectroscopy. The average ± SD value from all cell types was D = 0.8 ± 0.2 μm2/s (n = 218; 25°C). This is threefold lower than the D in blebs formed on Rat1 cells, D = 2.5 ± 0.8 μm2/s (n = 26). It is also significantly lower than the D in the outer leaflet or in giant unilamellar vesicles and the diffusion coefficient for other lipids on the inner leaflet of these cell membranes. The simplest interpretation is that approximately two thirds of the PIP2 on inner leaflet of these plasma membranes is bound reversibly.

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