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 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 Control of dynamic cell behaviors during angiogenesis and anastomosis by Rasip 1

Development ◽  
2021 ◽  
Author(s):  
Minkyoung Lee ◽  
Charles Betz ◽  
Jianmin Yin ◽  
Ilkka Paatero ◽  
Niels Schellinx ◽  
...  
Keyword(s):  
Cell Junctions ◽  
Interacting Protein ◽  
Cell Behaviors ◽  
Sprouting Angiogenesis ◽  
Junction Formation ◽  
Phenotype Analysis ◽  
Membrane Compartment ◽  
Dynamic Cell ◽  
Cellular Behaviors ◽  
And Function

Organ morphogenesis is driven by a wealth of tightly orchestrated cellular behaviors, which ensure proper organ assembly and function. Many of these cell activities involve cell-cell interactions and remodeling of the F-actin cytoskeleton. Here, we analyze the requirement for Rasip1 (Ras-interacting protein 1), an endothelial-specific regulator of junctional dynamics, during blood vessel formation. Phenotype analysis of rasip1 mutants in zebrafish embryos reveal distinct functions of Rasip1 during sprouting angiogenesis, anastomosis and lumen formation. During angiogenic sprouting, loss of Rasip1 causes cell pairing defects due to a destabilization of tricellular junctions, indicating that stable tri-cellular junctions are essential to maintain multicellular organization within the sprout. During anastomosis, Rasip1 is required to establish a stable apical membrane compartment; rasip1 mutants display ectopic, reticulated junctions and the apical compartment is frequently collapsed. Loss of Ccm1 and Heg1 function mimics junctional defects of rasip1 mutants. Furthermore, downregulation of ccm1 and heg1 leads to a delocalization of Rasip1 at cell junctions, indicating that junctional tethering of Rasip1 is required for its function during junction formation and stabilization during sprouting angiogenesis.

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 Excess centrosomes disrupt vascular lumenization and endothelial cell adherens junctions

Angiogenesis ◽  
2020 ◽  
Vol 23 (4) ◽  
pp. 567-575
Author(s):  
Danielle B. Buglak ◽  
Erich J. Kushner ◽  
Allison P. Marvin ◽  
Katy L. Davis ◽  
Victoria L. Bautch
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Polarity ◽  
Adherens Junctions ◽  
Cell Junctions ◽  
Microtubule Organizing Center ◽  
Sprouting Angiogenesis ◽  
Organizing Center ◽  
Important Regulator ◽  
Cell Cell

Abstract Proper blood vessel formation requires coordinated changes in endothelial cell polarity and rearrangement of cell–cell junctions to form a functional lumen. One important regulator of cell polarity is the centrosome, which acts as a microtubule organizing center. Excess centrosomes perturb aspects of endothelial cell polarity linked to migration, but whether centrosome number influences apical–basal polarity and cell–cell junctions is unknown. Here, we show that excess centrosomes alter the apical–basal polarity of endothelial cells in angiogenic sprouts and disrupt endothelial cell–cell adherens junctions. Endothelial cells with excess centrosomes had narrower lumens in a 3D sprouting angiogenesis model, and zebrafish intersegmental vessels had reduced perfusion following centrosome overduplication. These results indicate that endothelial cell centrosome number regulates proper lumenization downstream of effects on apical–basal polarity and cell–cell junctions. Endothelial cells with excess centrosomes are prevalent in tumor vessels, suggesting how centrosomes may contribute to tumor vessel dysfunction.

scholarly journals Septin Roles and Mechanisms in Organization of Endothelial Cell Junctions

2020 ◽  
Author(s):  
Joanna Kim ◽  
John A. Cooper
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Adherens Junctions ◽  
Cell Junctions ◽  
Cell Junction ◽  
Cell Contact ◽  
Junction Molecules ◽  
Tnf Α ◽  
Multiple Cell ◽  
Cell Cell

AbstractSeptins play an important role in regulating the barrier function of the endothelial monolayer of the microvasculature. Depletion of septin 2 protein alters the organization of vascular endothelial (VE)-cadherin at cell-cell adherens junctions as well as the dynamics of membrane protrusions at endothelial cell-cell contact sites. Here, we report the discovery that localization of septin 2 at endothelial cell junctions is important for the distribution of a number of other junctional molecules. We also found that treatment of microvascular endothelial cells with the inflammatory mediator TNF-α led to sequestration of septin 2 away from cell junctions and into the cytoplasm, without an effect on the overall level of septin 2 protein. Interestingly, TNF-α treatment of endothelial monolayers produced effects similar to those of depletion of septin 2 on various molecular components of adherens junctions (AJs) and tight junctions (TJs). Immunofluorescence staining revealed disruption of the integrity of AJs and TJs at cell-cell junctions without significant changes in protein expression except for VE-cadherin and nectin-2. To investigate the mechanism of junctional localization of septin 2, we mutated the polybasic motif of septin 2, which is proposed to interact with PIP2 in the plasma membrane. Overexpression of PIP2-binding mutant (PIP2BM) septin 2 led to loss of septin 2 from cell junctions with accumulation in the cytoplasm. This redistribution of septin 2 away from the membrane led to effects on cell junction molecules similar to those observed for depletion of septin 2. We conclude that septin localization to the membrane is essential for function and that septins support the localization of multiple cell junction molecules in endothelial cells.

scholarly journals Author response: Heart of glass anchors Rasip1 at endothelial cell-cell junctions to support vascular integrity

2015 ◽  
Author(s):  
Bart-Jan de Kreuk ◽  
Alexandre R Gingras ◽  
James DR Knight ◽  
Jian J Liu ◽  
Anne-Claude Gingras ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Cell Junctions ◽  
Author Response ◽  
Vascular Integrity ◽  
Cell Cell

scholarly journals Heart of glass anchors Rasip1 at endothelial cell-cell junctions to support vascular integrity

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Bart-Jan de Kreuk ◽  
Alexandre R Gingras ◽  
James DR Knight ◽  
Jian J Liu ◽  
Anne-Claude Gingras ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Binding Site ◽  
Central Region ◽  
Cell Junctions ◽  
Cardiovascular Development ◽  
Vascular Integrity ◽  
Proteomic Screen ◽  
Sequence Deletion ◽  
Rock Activity ◽  
Cell Cell

Heart of Glass (HEG1), a transmembrane receptor, and Rasip1, an endothelial-specific Rap1-binding protein, are both essential for cardiovascular development. Here we performed a proteomic screen for novel HEG1 interactors and report that HEG1 binds directly to Rasip1. Rasip1 localizes to forming endothelial cell (EC) cell-cell junctions and silencing HEG1 prevents this localization. Conversely, mitochondria-targeted HEG1 relocalizes Rasip1 to mitochondria in cells. The Rasip1-binding site in HEG1 contains a 9 residue sequence, deletion of which abrogates HEG1’s ability to recruit Rasip1. HEG1 binds to a central region of Rasip1 and deletion of this domain eliminates Rasip1’s ability to bind HEG1, to translocate to EC junctions, to inhibit ROCK activity, and to maintain EC junctional integrity. These studies establish that the binding of HEG1 to Rasip1 mediates Rap1-dependent recruitment of Rasip1 to and stabilization of EC cell-cell junctions.

scholarly journals Excess Centrosomes Disrupt Vascular Lumenization and Endothelial Cell Adherens Junctions

2019 ◽  
Author(s):  
Danielle B Buglak ◽  
Erich J Kushner ◽  
Allison P Marvin ◽  
Katy L Davis ◽  
Victoria L Bautch
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Polarity ◽  
Adherens Junctions ◽  
Cell Junctions ◽  
Microtubule Organizing Center ◽  
Sprouting Angiogenesis ◽  
Organizing Center ◽  
Important Regulator ◽  
Cell Cell

ABSTRACTProper blood vessel formation requires coordinated changes in endothelial cell polarity and rearrangement of cell-cell junctions to form a functional lumen. One important regulator of cell polarity is the centrosome, which acts as a microtubule organizing center. Excess centrosomes perturb aspects of endothelial cell polarity linked to migration, but whether centrosome number influences apical-basal polarity and cell-cell junctions is unknown. Here, we show that excess centrosomes alter the apical-basal polarity of endothelial cells in angiogenic sprouts and disrupt endothelial cell-cell adherens junctions. Endothelial cells with excess centrosomes had narrower lumens in a 3D sprouting angiogenesis model, and zebrafish intersegmental vessels had reduced perfusion following centrosome overduplication. These results indicate that endothelial cell centrosome number regulates proper lumenization downstream of effects on apical-basal polarity and cell-cell junctions. Endothelial cells with excess centrosomes are prevalent in tumor vessels, suggesting how centrosomes may contribute to tumor vessel dysfunction.

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.

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