scholarly journals A mechanosensing mechanism mediated by IRSp53 controls plasma membrane shape homeostasis at the nanoscale

2021 ◽  
Author(s):  
Xarxa Quiroga ◽  
Nikhil Walani ◽  
Albert Chavero ◽  
Alexandra Mittens ◽  
Andrea Disanza ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Feedback Loop ◽  
Actin Polymerization ◽  
Nano Scale ◽  
Mechanical Deformations ◽  
Cell Stretch ◽  
Potential Applicability ◽  
Membrane Shape

As cells migrate and experience forces from their surroundings, they constantly undergo mechanical deformations which reshape their plasma membrane (PM). To maintain homeostasis, cells need to detect and restore such changes, not only in terms of overall PM area and tension as previously described, but also in terms of local, nano-scale topography. Here we describe a novel phenomenon, by which cells sense and restore mechanically induced PM nano-scale deformations. We show that cell stretch and subsequent compression reshape the PM in a way that generates local membrane evaginations in the 100 nm scale. These evaginations are recognized by the I-BAR protein IRSp53, which triggers a burst of actin polymerization mediated by Rac1 and Arp2/3. The actin polymerization burst subsequently re-flattens the evagination, completing the mechanochemical feedback loop. Our results demonstrate a new mechanosensing mechanism for PM shape homeostasis, with potential applicability in different physiological scenarios.

scholarly journals Visualizing the functional architecture of the endocytic machinery

eLife ◽  
2015 ◽  
Vol 4 ◽  
Author(s):  
Andrea Picco ◽  
Markus Mund ◽  
Jonas Ries ◽  
François Nédélec ◽  
Marko Kaksonen
Keyword(s):  
Plasma Membrane ◽  
Actin Polymerization ◽  
Coat Proteins ◽  
Superresolution Imaging ◽  
Dynamic Architecture ◽  
Endocytic Protein ◽  
Endocytic Machinery ◽  
Microscopy Method ◽  
Protein Components ◽  
Membrane Shape

Clathrin-mediated endocytosis is an essential process that forms vesicles from the plasma membrane. Although most of the protein components of the endocytic protein machinery have been thoroughly characterized, their organization at the endocytic site is poorly understood. We developed a fluorescence microscopy method to track the average positions of yeast endocytic proteins in relation to each other with a time precision below 1 s and with a spatial precision of ∼10 nm. With these data, integrated with shapes of endocytic membrane intermediates and with superresolution imaging, we could visualize the dynamic architecture of the endocytic machinery. We showed how different coat proteins are distributed within the coat structure and how the assembly dynamics of N-BAR proteins relate to membrane shape changes. Moreover, we found that the region of actin polymerization is located at the base of the endocytic invagination, with the growing ends of filaments pointing toward the plasma membrane.

scholarly journals Using kICS to Reveal Changed Membrane Diffusion of AQP-9 Treated with Drugs

Membranes ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 568
Author(s):  
Jakob L. Kure ◽  
Thommie Karlsson ◽  
Camilla B. Andersen ◽  
B. Christoffer Lagerholm ◽  
Vesa Loitto ◽  
...  
Keyword(s):  
Diffusion Coefficient ◽  
Plasma Membrane ◽  
Membrane Proteins ◽  
Actin Polymerization ◽  
Correlation Spectroscopy ◽  
Image Correlation ◽  
Membrane Diffusion ◽  
Hek 293 ◽  
293 Cells ◽  
Aquaporin 9

The formation of nanodomains in the plasma membrane are thought to be part of membrane proteins regulation and signaling. Plasma membrane proteins are often investigated by analyzing the lateral mobility. k-space ICS (kICS) is a powerful image correlation spectroscopy (ICS) technique and a valuable supplement to fluorescence correlation spectroscopy (FCS). Here, we study the diffusion of aquaporin-9 (AQP9) in the plasma membrane, and the effect of different membrane and cytoskeleton affecting drugs, and therefore nanodomain perturbing, using kICS. We measured the diffusion coefficient of AQP9 after addition of these drugs using live cell Total Internal Reflection Fluorescence imaging on HEK-293 cells. The actin polymerization inhibitors Cytochalasin D and Latrunculin A do not affect the diffusion coefficient of AQP9. Methyl-β-Cyclodextrin decreases GFP-AQP9 diffusion coefficient in the plasma membrane. Human epidermal growth factor led to an increase in the diffusion coefficient of AQP9. These findings led to the conclusion that kICS can be used to measure diffusion AQP9, and suggests that the AQP9 is not part of nanodomains.

Coupled sterol synthesis and transport machineries at ER–endocytic contact sites

2021 ◽  
Vol 220 (10) ◽  
Author(s):  
Javier Encinar del Dedo ◽  
Isabel María Fernández-Golbano ◽  
Laura Pastor ◽  
Paula Meler ◽  
Cristina Ferrer-Orta ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Plasma Membrane ◽  
Actin Polymerization ◽  
Sterol Synthesis ◽  
Biosynthetic Enzymes ◽  
Cellular Membranes ◽  
Contact Sites ◽  
Endocytic Machinery ◽  
Mother Cells

Sterols are unevenly distributed within cellular membranes. How their biosynthetic and transport machineries are organized to generate heterogeneity is largely unknown. We previously showed that the yeast sterol transporter Osh2 is recruited to endoplasmic reticulum (ER)–endocytic contacts to facilitate actin polymerization. We now find that a subset of sterol biosynthetic enzymes also localizes at these contacts and interacts with Osh2 and the endocytic machinery. Following the sterol dynamics, we show that Osh2 extracts sterols from these subdomains, which we name ERSESs (ER sterol exit sites). Further, we demonstrate that coupling of the sterol synthesis and transport machineries is required for endocytosis in mother cells, but not in daughters, where plasma membrane loading with accessible sterols and endocytosis are linked to secretion.

scholarly journals More than a mere supply of monomers: G-Actin pools regulate actin dynamics in dendritic spines

2017 ◽  
Vol 216 (8) ◽  
pp. 2255-2257 ◽  
Author(s):  
Katalin Schlett
Keyword(s):  
Plasma Membrane ◽  
Dendritic Spines ◽  
Actin Polymerization ◽  
Synaptic Activity ◽  
Actin Dynamics ◽  
Actin Monomer ◽  
Cell Biol ◽  
Local Enrichment

Synaptic activity reshapes the morphology of dendritic spines via regulating F-actin arborization. In this issue, Lei et al. (2017. J. Cell Biol. https://doi.org/10.1083/jcb.201612042) reports a novel, G-actin–dependent regulation of actin polymerization within spine heads. They show that actin monomer levels are elevated in spines upon activity, with G-actin immobilized by the local enrichment of phosphatidylinositol (3,4,5)-triphosphate (PIP3) within the spine plasma membrane.

Activation of the small GTP-binding proteins rho and rac by growth factor receptors

1995 ◽  
Vol 108 (1) ◽  
pp. 225-233 ◽  
Author(s):  
C.D. Nobes ◽  
P. Hawkins ◽  
L. Stephens ◽  
A. Hall
Keyword(s):  
Plasma Membrane ◽  
Growth Factor ◽  
Tyrosine Kinase ◽  
Phorbol Ester ◽  
Lysophosphatidic Acid ◽  
Actin Polymerization ◽  
Binding Proteins ◽  
Platelet Derived Growth Factor ◽  
Gtp Binding ◽  
Rho Protein

The small GTP-binding proteins, rho and rac, control signal transduction pathways that link growth factor receptors to the activation of actin polymerization. In Swiss 3T3 cells, rho proteins mediate the lysophosphatidic acid and bombesin-induced formation of focal adhesions and actin stress fibres, whilst rac proteins are required for the platelet-derived growth factor-, insulin-, bombesin- and phorbol ester (phorbol 12-myristate 13-acetate)-stimulated actin polymerization at the plasma membrane that results in membrane ruffling. To investigate the role of p85/p110 phosphatidylinositol 3-kinase in the rho and rac signalling pathways, we have used a potent inhibitor of this activity, wortmannin. Wortmannin has no effect on focal adhesion or actin stress fibre formation induced by lysophosphatidic acid, bombesin or microinjected recombinant rho protein. In contrast, it totally inhibits plasma membrane edge-ruffling induced by platelet-derived growth factor and insulin though not by bombesin, phorbol ester or microinjected recombinant rac protein. We conclude that phosphatidylinositol 3,4,5 trisphosphate mediates activation of rac by the platelet-derived growth factor and insulin receptors. The effects of lysophosphatidic acid on the Swiss 3T3 actin cytoskeleton can be blocked by the tyrosine kinase inhibitor, tyrphostin. Since tyrphostin does not inhibit the effects of microinjected rho protein, we conclude that lysophosphatidic acid activation of rho is mediated by a tyrosine kinase.

scholarly journals Actin and microtubule cross talk mediates persistent polarized growth

2018 ◽  
Vol 217 (10) ◽  
pp. 3531-3544 ◽  
Author(s):  
Shu-Zon Wu ◽  
Magdalena Bezanilla
Keyword(s):  
Cell Expansion ◽  
Feedback Loop ◽  
Actin Polymerization ◽  
Physcomitrella Patens ◽  
Polarized Growth ◽  
Cellular Processes ◽  
Functional Connections ◽  
Class Viii ◽  
Direct Localization ◽  
Open Question

Coordination between actin and microtubules is important for numerous cellular processes in diverse eukaryotes. In plants, tip-growing cells require actin for cell expansion and microtubules for orientation of cell expansion, but how the two cytoskeletons are linked is an open question. In tip-growing cells of the moss Physcomitrella patens, we show that an actin cluster near the cell apex dictates the direction of rapid cell expansion. Formation of this structure depends on the convergence of microtubules near the cell tip. We discovered that microtubule convergence requires class VIII myosin function, and actin is necessary for myosin VIII–mediated focusing of microtubules. The loss of myosin VIII function affects both networks, indicating functional connections among the three cytoskeletal components. Our data suggest that microtubules direct localization of formins, actin nucleation factors, that generate actin filaments further focusing microtubules, thereby establishing a positive feedback loop ensuring that actin polymerization and cell expansion occur at a defined site resulting in persistent polarized growth.

scholarly journals Requirement for Formin-Induced Actin Polymerization during Spread of Shigella flexneri

2009 ◽  
Vol 78 (1) ◽  
pp. 193-203 ◽  
Author(s):  
Jason E. Heindl ◽  
Indrani Saran ◽  
Chae-ryun Yi ◽  
Cammie F. Lesser ◽  
Marcia B. Goldberg
Keyword(s):  
Plasma Membrane ◽  
Virus Infection ◽  
Vaccinia Virus ◽  
Cell Body ◽  
Actin Polymerization ◽  
Shigella Flexneri ◽  
Microbial Pathogens ◽  
Bacterial Surface ◽  
Virulence Protein ◽  
Intercellular Spread

ABSTRACT Actin polymerization in the cytosol and at the plasma membrane is locally regulated by actin nucleators. Several microbial pathogens exploit cellular actin polymerization to spread through tissue. The movement of the enteric pathogen Shigella flexneri, both within the cell body and from cell to cell, depends on actin polymerization. During intercellular spread, actin polymerization at the bacterial surface generates protrusions of the plasma membrane, which are engulfed by adjacent cells. In the cell body, polymerization of actin by Shigella spp. is dependent on N-WASP activation of the Arp2/Arp3 complex. Here we demonstrate that, in contrast, efficient protrusion formation and intercellular spread depend on actin polymerization that involves activation of the Diaphanous formin Dia. While the Shigella virulence protein IpgB2 can bind and activate Dia1 (N. M. Alto et al., Cell 124:133-145, 2006), its absence does not result in a detectable defect in Dia-dependent protrusion formation or spread. The dependence on the activation of Dia during S. flexneri infection contrasts with the inhibition of this pathway observed during vaccinia virus infection.

scholarly journals TCR–pMHC bond conformation controls TCR ligand discrimination

2019 ◽  
Vol 17 (3) ◽  
pp. 203-217 ◽  
Author(s):  
Dibyendu K. Sasmal ◽  
Wei Feng ◽  
Sobhan Roy ◽  
Peter Leung ◽  
Yanran He ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Energy Transfer ◽  
Feedback Loop ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Intrinsic Property ◽  
Unanswered Question ◽  
Structural Differences ◽  
Self Peptides

Abstract A major unanswered question is how a TCR discriminates between foreign and self-peptides presented on the APC surface. Here, we used in situ fluorescence resonance energy transfer (FRET) to measure the distances of single TCR–pMHC bonds and the conformations of individual TCR–CD3ζ receptors at the membranes of live primary T cells. We found that a TCR discriminates between closely related peptides by forming single TCR–pMHC bonds with different conformations, and the most potent pMHC forms the shortest bond. The bond conformation is an intrinsic property that is independent of the binding affinity and kinetics, TCR microcluster formation, and CD4 binding. The bond conformation dictates the degree of CD3ζ dissociation from the inner leaflet of the plasma membrane via a positive calcium signaling feedback loop to precisely control the accessibility of CD3ζ ITAMs for phosphorylation. Our data revealed the mechanism by which a TCR deciphers the structural differences among peptides via the TCR–pMHC bond conformation.

scholarly journals Neph1 Cooperates with Nephrin To Transduce a Signal That Induces Actin Polymerization

2007 ◽  
Vol 27 (24) ◽  
pp. 8698-8712 ◽  
Author(s):  
Puneet Garg ◽  
Rakesh Verma ◽  
Deepak Nihalani ◽  
Duncan B. Johnstone ◽  
Lawrence B. Holzman
Keyword(s):  
Plasma Membrane ◽  
Actin Polymerization ◽  
Intercellular Junction ◽  
Actin Dynamics ◽  
Cellular Motility ◽  
Tyrosine Residues ◽  
Signaling Complex ◽  
Direct Assembly ◽  
Junction Formation ◽  
Unique Model

ABSTRACT While the mechanisms that regulate actin dynamics in cellular motility are intensively studied, relatively little is known about signaling events that transmit outside-in signals and direct assembly and regulation of actin polymerization complexes at the cell membrane. The kidney podocyte provides a unique model for investigating these mechanisms since deletion of Nephrin or Neph1, two interacting components of the specialized podocyte intercellular junction, results in abnormal podocyte morphogenesis and junction formation. We provide evidence that extends the existing model by which the Nephrin-Neph1 complex transduces phosphorylation-mediated signals that assemble an actin polymerization complex at the podocyte intercellular junction. Upon engagement, Neph1 is phosphorylated on specific tyrosine residues by Fyn, which results in the recruitment of Grb2, an event that is necessary for Neph1-induced actin polymerization at the plasma membrane. Importantly, Neph1 and Nephrin directly interact and, by juxtaposing Grb2 and Nck1/2 at the membrane following complex activation, cooperate to augment the efficiency of actin polymerization. These data provide evidence for a mechanism reminiscent of that employed by vaccinia virus and other pathogens, by which a signaling complex transduces an outside-in signal that results in actin filament polymerization at the plasma membrane.

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