Biology subjects

Huveneers, S.

Publications and source records attributed to Huveneers, S..

7 recordsLinked to original sources

bioRxiv2025

Phosphoproteomics Maps Calcineurin-NFAT-DSCR1.4 Signaling as Druggable Axis in Gαq-R183Q-Driven Capillary Malformations

Vascular malformations are congenital lesions caused by somatic and germline mutations that disrupt developmental signaling pathways. Capillary malformations (CMs) typically present as port-wine stains in the skin and can also affect ocular and cerebral tissues in Sturge Weber Syndrome (SWS), leading to aesthetic, ophthalmic, and neurological complications. CMs are caused by a somatic mutation in the GNAQ gene in endothelial cells, leading to a p.R183Q substitution in the Gq protein. The underlying mechanisms of Gq-R183Q-driven CMs formation remain unclear. To address this, we generated CRISPR/Cas9-engineered human dermal microvascular endothelial cells lacking endogenous Gq, whilst expressing the Gq-R183Q mutant instead. The Gq-R183Q mutation strongly impaired endothelial cell migration and angiogenic sprouting capacity compared to wild-type controls. Next, using SILAC-based quantitative proteomics, we investigated the Gq-R183Q-induced changes in the endothelial phosphoproteome. These analyses revealed prominent activation of the calcineurin-NFAT signaling pathway in Gq-R183Q-expressing endothelial cells, leading to dephosphorylation of NFAT1 and NFAT2 and the selective expression of their transcriptional target DSCR1.4. Immunofluorescence of patient-derived skin biopsies confirmed deregulation of NFAT1/2 and the expression of DSRC1 in endothelial cells, validating their potential importance in CMs. We further demonstrate that pharmacological inhibition of calcineurin with tacrolimus (FK506) could partially restore NFAT signaling in Gq-R183Q endothelial cells. Intriguingly, the genetic depletion of the NFAT target DSCR1 in Gq-R183Q cells fully restored calcineurin/NFAT signaling to normal levels, enabling proper endothelial migration and sprouting. In summary, we uncovered a calcineurin-NFAT-DSCR1.4 signal transduction axis that is driven by Gq-R183Q and established its importance for endothelial angiogenic properties. These findings highlight the calcineurin/NFAT signaling axis as a promising therapeutic target to restore endothelial function in CMs.

cell biology↗

bioRxiv2025

VE-cadherin RGD motifs are dispensable for cell-cell junctions, endothelial barrier function and leukocyte extravasation

VE-cadherin is a key transmembrane protein in endothelial cell-cell junctions, essential for maintaining vascular integrity and regulating selective leukocyte extravasation into inflamed tissue. The extracellular domain of human VE-cadherin contains two arginine-glycine-aspartate (RGD) motifs, which are known integrin-binding sites, particularly for integrins in the {beta}1, {beta}3, and {beta}5 families. In this study, we examined the functional relevance of these RGD motifs by generating VE-cadherin variants with the RGD sequences mutated to non-functional RGE. Immunofluorescence analysis showed that the VE-cadherin[D238E], VE-cadherin[D301E], and double-mutant VE-cadherin[D238/301E] variants formed stable cell-cell junctions, comparable to wild-type VE-cadherin. Additionally, electric cell-substrate impedance sensing (ECIS) confirmed that endothelial cells expressing each VE-cadherin RGD>RGE variant maintained efficient barrier function. Moreover, leukocyte transmigration assays demonstrated that the RGD>RGE mutations did not affect leukocyte-endothelial interactions during transmigration. In summary, our findings indicate that the VE-cadherin RGD motifs are not essential for endothelial junction formation or leukocyte transmigration.

cell biology↗

bioRxiv2024

Tumor-induced endothelial RhoA activation mediates tumor cell transendothelial migration and metastasis

Abstract/SummaryThe endothelial barrier plays an active role in transendothelial tumor cell migration during metastasis, however, the endothelial regulatory elements of this step remain obscure. Here we show that endothelial RhoA activation is a determining factor during this process. Breast tumor cell-induced endothelial RhoA activation is the combined outcome of paracrine IL-8-dependent and cell-to-cell contact {beta}1 integrin-mediated mechanisms, with elements of this pathway correlating with clinical data. Endothelial-specific RhoA blockade or in vivo deficiency inhibited the transendothelial migration and metastatic potential of human breast tumor and three murine syngeneic tumor cell lines, similar to the pharmacological blockade of the downstream RhoA pathway. These findings highlight endothelial RhoA as a potent, universal target in the tumor microenvironment for anti-metastatic treatment of solid tumors.

physiology↗

bioRxiv2024

Ultrasound-activated microbubbles mediate F-actin disruptions and endothelial gap formation during sonoporation

Locally opening up the endothelial barrier in a safe and controlled way is beneficial for drug delivery into the extravascular tissue. Although ultrasound-induced microbubble oscillations can affect endothelial barrier integrity, the mechanism remains unknown. Here we uncover a new role for F-actin in microbubble-mediated endothelial gap formation. Unique simultaneous high-resolution confocal microscopy and ultra-high-speed camera imaging (10 million frames per second) reveal that oscillating microbubbles (radius 1.3-3.8 {micro}m) induce sonoporation in all cells in which F-actin remodeling occurred. F-actin disruption only mainly resulted in tunnel formation (75%) and F-actin stress fiber severing and recoil mainly resulted in cell-cell contact opening within 15 s upon treatment (54%) and tunnel formation (15%). Stress fiber severing occurred when fibers were within reach of the microbubbles maximum radius during oscillation, requiring normal forces of [≥]230 nN. Together, these findings reveal a novel mechanism of microbubble-mediated drug delivery, which associates with the underlying cytoskeletal organization.

bioengineering↗

bioRxiv2021

Vinculin controls endothelial cell junction dynamics during vascular lumen formation

Blood 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.

cell biology↗

bioRxiv2019

DLC1 is a direct target of activated YAP/TAZ that drives collective migration and sprouting angiogenesis

YAP/TAZ signaling is crucial for sprouting angiogenesis and vascular homeostasis through the regulation of endothelial remodeling. Thus far the underlying molecular mechanisms that explain how YAP/TAZ control the vasculature remain unclear. We here identify Deleted-in-Liver-Cancer-1 (DLC1) as a direct transcriptional target of the activated YAP/TAZ-TEAD complex in the endothelium. Substrate stiffening and VEGF stimuli promote the endothelial expression of DLC1. DLC1 expression is dependent on the presence of YAP and TAZ, and constitutive activation of YAP efficiently promotes expression of DLC1. We show that DLC1 limits F-actin fiber formation, integrin-based focal adhesion lifetime and integrin-mediated traction forces. Depletion of endothelial DLC1 strongly perturbs cell polarization in directed collective migration and inhibits the formation of angiogenic sprouts. Importantly, the inability of YAP-depleted endothelial cells to collectively migrate and form angiogenic sprouts can be rescued by ectopic expression of DLC1. Together, these findings reveal that DLC1 fills a hitherto missing link between YAP/TAZ signaling and endothelial dynamics during angiogenesis.

cell biology↗

bioRxiv2018

Hematopoietic Stem And Progenitor Cells Use Podosomes To Transcellularly Home To The Bone Marrow.

Bone marrow (BM) endothelium plays an important role in homing of hematopoietic stem and progenitor cells (HSPCs) upon transplantation, but surprisingly little is known on how endothelial cells regulate local permeability and HSPC transmigration. We show that temporal loss of VE-cadherin function promotes vascular permeability in BM, even upon low dose irradiation and strongly enhanced homing of transplanted HSPCs to BM of irradiated mice. Intriguingly, stabilizing junctional VE-cadherin in vivo reduced BM permeability, but did not prevent HSPC migration into the BM, suggesting that HSPCs enter the bone marrow by transcellularly crossing the endothelium. Indeed, HSPCs induce podosomes to cross human BM endothelial monolayers in a transcellular manner. By contrast, HSPC rather use the paracellular route when VE-cadherin function is inhibited. Taken together, VE-cadherin is crucial for BM vascular homeostasis and HSPC homing, and may therefore serve as a potential therapeutic target to improve HSPC homing strategies.

cell biology↗