scholarly journals Golgi-associated cPLA2α Regulates Endothelial Cell–Cell Junction Integrity by Controlling the Trafficking of Transmembrane Junction Proteins

2009 ◽  
Vol 20 (19) ◽  
pp. 4225-4234 ◽  
Author(s):  
Elsa Regan-Klapisz ◽  
Vincent Krouwer ◽  
Miriam Langelaar-Makkinje ◽  
Laxman Nallan ◽  
Michael Gelb ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Intracellular Trafficking ◽  
Adherens Junction ◽  
Cell Junctions ◽  
Cell Junction ◽  
Tight Junction Formation ◽  
Junction Formation ◽  
Junction Proteins ◽  
Cell Cell

In endothelial cells specifically, cPLA2α translocates from the cytoplasm to the Golgi complex in response to cell confluence. Considering the link between confluence and cell–cell junction formation, and the emerging role of cPLA2α in intracellular trafficking, we tested whether Golgi-associated cPLA2α is involved in the trafficking of junction proteins. Here, we show that the redistribution of cPLA2α from the cytoplasm to the Golgi correlates with adherens junction maturation and occurs before tight junction formation. Disruption of adherens junctions using a blocking anti-VE-cadherin antibody reverses the association of cPLA2α with the Golgi. Silencing of cPLA2α and inhibition of cPLA2α enzymatic activity using various inhibitors result in the diminished presence of the transmembrane junction proteins VE-cadherin, occludin, and claudin-5 at cell–cell contacts, and in their accumulation at the Golgi. Altogether, our data support the idea that VE-cadherin triggers the relocation of cPLA2α to the Golgi and that in turn, Golgi-associated cPLA2α regulates the transport of transmembrane junction proteins through or from the Golgi, thereby controlling the integrity of endothelial cell–cell junctions.

scholarly journals PKM2 regulates endothelial cell junction dynamics and angiogenesis via ATP production

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Jesús Gómez-Escudero ◽  
Cristina Clemente ◽  
Diego García-Weber ◽  
Rebeca Acín-Pérez ◽  
Jaime Millán ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Inflammatory Disease ◽  
Chronic Inflammatory Disease ◽  
Cell Junctions ◽  
Cell Junction ◽  
Collective Migration ◽  
Sprouting Angiogenesis ◽  
Cell Cell

Abstract Angiogenesis, the formation of new blood vessels from pre-existing ones, occurs in pathophysiological contexts such as wound healing, cancer, and chronic inflammatory disease. During sprouting angiogenesis, endothelial tip and stalk cells coordinately remodel their cell-cell junctions to allow collective migration and extension of the sprout while maintaining barrier integrity. All these processes require energy, and the predominant ATP generation route in endothelial cells is glycolysis. However, it remains unclear how ATP reaches the plasma membrane and intercellular junctions. In this study, we demonstrate that the glycolytic enzyme pyruvate kinase 2 (PKM2) is required for sprouting angiogenesis in vitro and in vivo through the regulation of endothelial cell-junction dynamics and collective migration. We show that PKM2-silencing decreases ATP required for proper VE-cadherin internalization/traffic at endothelial cell-cell junctions. Our study provides fresh insight into the role of ATP subcellular compartmentalization in endothelial cells during angiogenesis. Since manipulation of EC glycolysis constitutes a potential therapeutic intervention route, particularly in tumors and chronic inflammatory disease, these findings may help to refine the targeting of endothelial glycolytic activity in disease.

scholarly journals Regulation of Adherens Junction Dynamics by Phosphorylation Switches

2012 ◽  
Vol 2012 ◽  
pp. 1-14 ◽  
Author(s):  
Cristina Bertocchi ◽  
Megha Vaman Rao ◽  
Ronen Zaidel-Bar
Keyword(s):  
Actin Cytoskeleton ◽  
Enzymatic Activity ◽  
Adherens Junctions ◽  
Adherens Junction ◽  
Adaptor Proteins ◽  
Cell Junction ◽  
Internal Forces ◽  
Adherens Junction Proteins ◽  
Junction Proteins

Adherens junctions connect the actin cytoskeleton of neighboring cells through transmembrane cadherin receptors and a network of adaptor proteins. The interactions between these adaptors and cadherin as well as the activity of actin regulators localized to adherens junctions are tightly controlled to facilitate cell junction assembly or disassembly in response to changes in external or internal forces and/or signaling. Phosphorylation of tyrosine, serine, or threonine residues acts as a switch on the majority of adherens junction proteins, turning “on” or “off” their interactions with other proteins and/or their enzymatic activity. Here, we provide an overview of the kinases and phosphatases regulating phosphorylation of adherens junction proteins and bring examples of phosphorylation events leading to the assembly or disassembly of adherens junctions, highlighting the important role of phosphorylation switches in regulating their dynamics.

scholarly journals KRIT-1/CCM1 is a Rap1 effector that regulates endothelial cell–cell junctions

2007 ◽  
Vol 179 (2) ◽  
pp. 247-254 ◽  
Author(s):  
Angela Glading ◽  
Jaewon Han ◽  
Rebecca A. Stockton ◽  
Mark H. Ginsberg
Keyword(s):  
Endothelial Cell ◽  
Protein Function ◽  
Cell Junctions ◽  
Stress Fibers ◽  
Ferm Domain ◽  
Endothelial Junctions ◽  
Guanosine Triphosphatase ◽  
Interfering Rna ◽  
Junction Proteins ◽  
Cell Cell

Cerebral cavernous malformation (CCM), a disease associated with defective endothelial junctions, result from autosomal dominant CCM1 mutations that cause loss of KRIT-1 protein function, though how the loss of KRIT-1 leads to CCM is obscure. KRIT-1 binds to Rap1, a guanosine triphosphatase that maintains the integrity of endothelial junctions. Here, we report that KRIT-1 protein is expressed in cultured arterial and venous endothelial cells and is present in cell–cell junctions. KRIT-1 colocalized and was physically associated with junctional proteins via its band 4.1/ezrin/radixin/moesin (FERM) domain. Rap1 activity regulated the junctional localization of KRIT-1 and its physical association with junction proteins. However, the association of the isolated KRIT-1 FERM domain was independent of Rap1. Small interfering RNA–mediated depletion of KRIT-1 blocked the ability of Rap1 to stabilize endothelial junctions associated with increased actin stress fibers. Thus, Rap1 increases KRIT-1 targeting to endothelial cell–cell junctions where it suppresses stress fibers and stabilizes junctional integrity.

scholarly journals Role of IQGAP1 in endothelial barrier enhancement caused by OxPAPC

2016 ◽  
Vol 311 (4) ◽  
pp. L800-L809 ◽  
Author(s):  
Yufeng Tian ◽  
Xinyong Tian ◽  
Grzegorz Gawlak ◽  
Nicolene Sarich ◽  
David B. Sacks ◽  
...  
Keyword(s):  
Actin Cytoskeleton ◽  
Actin Polymerization ◽  
Adherens Junctions ◽  
Adherens Junction ◽  
Cell Junctions ◽  
Endothelial Barrier ◽  
Cell Junction ◽  
Protective Effects ◽  
P120 Catenin

Oxidized 1-palmitoyl-2-arachidonoyl- sn-glycero-3-phosphatidylcholine (OxPAPC) attenuates agonist-induced endothelial cell (EC) permeability and increases pulmonary endothelial barrier function via enhancement of both the peripheral actin cytoskeleton and cell junctions mediated by Rac1 and Cdc42 GTPases. This study evaluated the role for the multifunctional Rac1/Cdc42 effector and regulator, IQ domain containing GTPase-activating protein (IQGAP1), as a molecular transducer of the OxPAPC-mediated EC barrier-enhancing signal. IQGAP1 knockdown in endothelial cells by gene-specific small-interfering RNA abolished OxPAPC-induced enlargement of VE-cadherin-positive adherens junctions, suppressed peripheral accumulation of actin polymerization regulators, namely cortactin, neural Wiskott-Aldrich syndrome protein (N-WASP), and actin-related protein 3, and attenuated remodeling of the peripheral actin cytoskeleton. Inhibition of OxPAPC-induced barrier enhancement by IQGAP1 knockdown was due to suppressed Rac1 and Cdc42 activation. Expression of an IQGAP1 truncated mutant showed that the GTPase regulatory domain of IQGAP1 was essential for the OxPAPC-induced membrane localization of cortactin, adherens junction proteins VE-cadherin and p120-catenin, as well as for EC permeability response. IQGAP1 knockdown attenuated the protective effect of OxPAPC against thrombin-induced cell contraction, cell junction disruption, and EC permeability. These results demonstrate for the first time the role of IQGAP1 as a critical transducer of OxPAPC-induced Rac1/Cdc42 signaling to the actin cytoskeleton and adherens junctions, which promotes cortical cytoskeletal remodeling and EC barrier-protective effects of oxidized phospholipids.

scholarly journals Vinculin controls endothelial cell junction dynamics during vascular lumen formation

2021 ◽  
Author(s):  
Maria P. Kotini ◽  
Miesje M. van der Stoel ◽  
Mitchell K. Han ◽  
Bettina Kirchmaier ◽  
Johan de Rooij ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Cell Junctions ◽  
Cell Junction ◽  
Vascular Integrity ◽  
Lumen Formation ◽  
Cell Behaviors ◽  
Sprouting Angiogenesis ◽  
Dynamic Cell ◽  
External Forces ◽  
Cell Cell

AbstractBlood vessel morphogenesis is driven by coordinated endothelial cell behaviors, which depend on dynamic cell-cell interactions. Remodeling of endothelial cell-cell junctions promote morphogenetic cellular events while preserving vascular integrity. Here, we have analyzed the dynamics of endothelial cell-cell junctions during lumen formation in angiogenic sprouts. By live-imaging of the formation of intersegmental blood vessels in zebrafish, we demonstrate that lumen expansion is accompanied by the formation of transient finger-shaped junctions. Formation and maintenance of these junctional fingers are positively regulated by blood pressure whereas inhibition of blood flow prevents their formation. Using fluorescent reporters, we show that the tension-sensor Vinculin localizes to junctional fingers. Furthermore, loss of vinculin function, in vinculin a and -b double knockouts, prevents junctional finger formation in angiogenic sprouts, whereas endothelial expression of a vinculin transgene is sufficient to restore junctional fingers. Taken together, our findings suggest a mechanism in which lumen expansion during angiogenesis leads to an increase in junctional tension, which triggers recruitment of vinculin and formation of junctional fingers. We propose that endothelial cells may employ force-dependent junctional remodeling to react to changes in external forces to protect cell-cell contacts and to maintain vascular integrity during sprouting angiogenesis.

scholarly journals PLEKHG4B enables actin cytoskeletal remodeling during epithelial cell-cell junction formation

2020 ◽  
pp. jcs.249078
Author(s):  
Komaki Ninomiya ◽  
Kai Ohta ◽  
Kazunari Yamashita ◽  
Kensaku Mizuno ◽  
Kazumasa Ohashi
Keyword(s):  
Epithelial Cell ◽  
A549 Cells ◽  
Cell Junctions ◽  
Cell Junction ◽  
Nucleotide Exchange ◽  
Actin Bundles ◽  
Rhoa Activity ◽  
Cytoskeletal Remodeling ◽  
Junction Formation ◽  
Cell Cell

Cell-cell junction formation requires actin cytoskeletal remodeling. Here we show that PLEKHG4B, a Rho-guanine nucleotide exchange factor (Rho-GEF), plays a crucial role in epithelial cell-cell junction formation. Knockdown of PLEKHG4B decreased Cdc42 activity and tended to increase RhoA activity in A549 cells. A549 monolayer cells showed 'closed junctions' with closely packed actin bundles along the cell-cell contacts, but PLEKHG4B knockdown suppressed closed junction formation and exhibited 'open junctions' with split actin bundles located away from the cell-cell boundary. In calcium-switch assays, PLEKHG4B knockdown delayed the conversion of open junctions to closed junctions and β-catenin accumulation at cell-cell junctions. Further, PLEKHG4B knockdown abrogated the reduction in myosin activity normally seen in the later stage of junction formation. The aberrant myosin activation and impairments in closed junction formation in PLEKHG4B-knockdown cells were reverted by ROCK inhibition or LARG/PDZ-RhoGEF knockdown. These results suggest that PLEKHG4B enables actin remodeling during epithelial cell-cell junction maturation, probably by reducing myosin activity in the later stage of junction formation, through suppressing LARG/PDZ-RhoGEF and RhoA-ROCK activities. We also showed that annexin-A2 participates in PLEKHG4B localization to cell-cell junctions.

scholarly journals Regulation of Endothelial Cell Motility by Complexes of Tetraspan Molecules CD81/TAPA-1 and CD151/PETA-3 with α3β1 Integrin Localized at Endothelial Lateral Junctions

1998 ◽  
Vol 141 (3) ◽  
pp. 791-804 ◽  
Author(s):  
María Yáñez-Mó ◽  
Arántzazu Alfranca ◽  
Carlos Cabañas ◽  
Mónica Marazuela ◽  
Reyes Tejedor ◽  
...  
Keyword(s):  
Wound Healing ◽  
Endothelial Cell ◽  
Cell Motility ◽  
Cell Polarization ◽  
Cell Junctions ◽  
Cell Junction ◽  
Cell Contact ◽  
Collagen Gels ◽  
Α3β1 Integrin ◽  
Cell Cell

Cell-to-cell junction structures play a key role in cell growth rate control and cell polarization. In endothelial cells (EC), these structures are also involved in regulation of vascular permeability and leukocyte extravasation. To identify novel components in EC intercellular junctions, mAbs against these cells were produced and selected using a morphological screening by immunofluorescence microscopy. Two novel mAbs, LIA1/1 and VJ1/16, specifically recognized a 25-kD protein that was selectively localized at cell–cell junctions of EC, both in the primary formation of cell monolayers and when EC reorganized in the process of wound healing. This antigen corresponded to the recently cloned platelet-endothelial tetraspan antigen CD151/PETA-3 (platelet-endothelial tetraspan antigen-3), and was consistently detected at EC cell–cell contact sites. In addition to CD151/PETA-3, two other members of the tetraspan superfamily, CD9 and CD81/ TAPA-1 (target of antiproliferative antibody-1), localized at endothelial cell-to-cell junctions. Biochemical analysis demonstrated molecular associations among tetraspan molecules themselves and those of CD151/ PETA-3 and CD9 with α3β1 integrin. Interestingly, mAbs directed to both CD151/PETA-3 and CD81/ TAPA-1 as well as mAb specific for α3 integrin, were able to inhibit the migration of ECs in the process of wound healing. The engagement of CD151/PETA-3 and CD81/TAPA-1 inhibited the movement of individual ECs, as determined by quantitative time-lapse video microscopy studies. Furthermore, mAbs against the CD151/PETA-3 molecule diminished the rate of EC invasion into collagen gels. In addition, these mAbs were able to increase the adhesion of EC to extracellular matrix proteins. Together these results indicate that CD81/TAPA-1 and CD151/PETA-3 tetraspan molecules are components of the endothelial lateral junctions implicated in the regulation of cell motility, either directly or by modulation of the function of the associated integrin heterodimers.

scholarly journals High Mobility Group Box 1 Mediates TMAO-Induced Endothelial Dysfunction

2019 ◽  
Vol 20 (14) ◽  
pp. 3570 ◽  
Author(s):  
Gurinder Bir Singh ◽  
Yang Zhang ◽  
Krishna M. Boini ◽  
Saisudha Koka
Keyword(s):  
Endothelial Cells ◽  
Endothelial Dysfunction ◽  
High Mobility ◽  
Prior Treatment ◽  
Cell Junctions ◽  
High Mobility Group ◽  
Cell Junction ◽  
Tlr4 Expression ◽  
Junction Proteins ◽  
Cell Cell

The intestinal microbe-derived metabolite trimethylamine N-oxide (TMAO) is implicated in the pathogenesis of cardiovascular diseases (CVDs). The molecular mechanisms of how TMAO induces atherosclerosis and CVDs’ progression are still unclear. In this regard, high-mobility group box protein 1 (HMGB1), an inflammatory mediator, has been reported to disrupt cell–cell junctions, resulting in vascular endothelial hyper permeability leading to endothelial dysfunction. The present study tested whether TMAO associated endothelial dysfunction results via HMGB1 activation. Biochemical and RT-PCR analysis showed that TMAO increased the HMGB1 expression in a dose-dependent manner in endothelial cells. However, prior treatment with glycyrrhizin, an HMGB1 binder, abolished the TMAO-induced HMGB1 production in endothelial cells. Furthermore, Western blot and immunofluorescent analysis showed significant decrease in the expression of cell–cell junction proteins ZO-2, Occludin, and VE-cadherin in TMAO treated endothelial cells compared with control cells. However, prior treatment with glycyrrhizin attenuated the TMAO-induced cell–cell junction proteins’ disruption. TMAO increased toll-like receptor 4 (TLR4) expression in endothelial cells. Inhibition of TLR4 expression by TLR4 siRNA protected the endothelial cells from TMAO associated tight junction protein disruption via HMGB1. In conclusion, our results demonstrate that HMGB1 is one of the important mediators of TMAO-induced endothelial dysfunction.

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