Biology subjects

Belting, H.-G.

Publications and source records attributed to Belting, H.-G..

6 recordsLinked to original sources

bioRxiv2025

An AmotL2 - Yap1 Module Integrates Flow and Junctional Mechanics to Specify Vascular Pruning Hotspots

Vascular networks constantly remodel to optimise blood flow, yet how endothelial cells (ECs) integrate haemodynamic cues with junctional mechanics to execute precise branch pruning remains unclear. Using live imaging and quantitative topology analysis in the zebrafish sub-intestinal vein plexus, we define the cellular events underlying flow-guided regression. Pruning occurs in geometrically disadvantaged, low-flow branches and is executed through transient polarity fluctuations, junctional remodelling and directed EC migration. We identify AmotL2 as an essential regulator of endothelial mechanical competence. AmotL2 maintains junctional architecture, actomyosin tension and appropriate cell packing, enabling ECs to polarise and rearrange in response to local flow differences. Loss of AmotL2 disrupts tension homeostasis, increases tissue compaction and prevents branch regression despite normal perfusion. In contrast, Yap1 plays an opposing, stabilising role, and its loss results in excessive pruning and network simplification. Together, AmotL2 and Yap1 constitute a mechanosensitive balancing module that interprets flow magnitude and vascular geometry to determine branch stability. This framework describes how mechanical forces sculpt vascular architecture during development and homeostasis.

developmental biology↗

bioRxiv2023

Genetic analysis of rab7 mutants in zebrafish

Vascular network formation requires the fusion of newly formed blood vessels and the emergence of a patent lumen between the newly established connections so that blood flow can start. Lumen formation has been shown to depend on the late endosomal/lysosomal pathway in various organs of animal tubular systems. Here, we identified a late endosomal/lysosomal vesicular fraction (Rab7/Lamp2) in early zebrafish angiogenic sprouts, which appears to contribute to apical membrane growth during lumen formation. To study the effect of the late endocytic pathway on vascular development, we generated mutant alleles for all three rab7 genes in zebrafish (rab7a, rab7ba, rab7bb). All rab7 genes are expressed in wild-type zebrafish and we did not detect any compensatory effects by the other rab7 isoforms in single knockout mutants, which were all viable. Only the triple mutant was lethal suggesting some functional redundancy. However, the different rab7 isoforms fulfil also at least partially independent functions because eggs laid from mothers lacking two rab7 (rab7a and/or rab7bb). showed reduced survival and contained enlarged yolk granules, suggesting maternal contribution of these two rab7. Finally, we observed minor effects on lumen formation in embryos which still express one copy of rab7. Our results support the notion that the late endocytic/lysosomal compartment contributes to lumen expansion.

genetics↗

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↗

bioRxiv2020

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

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 requirements for Rasip1 during sprouting angiogenesis, vascular anastomosis and lumen formation. During angiogenic sprouting, Rasip1 is required for efficient cell pairing, which is essential for multicellular tube formation. High-resolution time-lapse analyses show that these cell pairing defects are caused by a destabilization of tricellular junctions suggesting that tri-cellular junctions may serve as a counterfort to tether sprouting endothelial cells during morphogenetic cell rearrangements. 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 leads to junctional defects similar to those seen in rasip1 mutants. Analysis of radil-b single and rasip1/radil-b double mutants reveal distinct and overlapping functions of both proteins. While Rasip1 and Radil-b have similar functions during angiogenic sprouting, the junction formation during anastomosis may primarily depend on Rasip1.

developmental biology↗

bioRxiv2018

Sprouting and anastomosis in the Drosophila trachea and the vertebrate vasculature: similarities and differences in cell behaviour

Branching morphogenesis is a fascinating process whereby a simple network of biological tubes increases its complexity by adding new branches to existing ones, generating an enlarged structure of interconnected tubes. Branching morphogenesis has been studied extensively in animals and much has been learned about the regulation of branching at the cellular and molecular level. Here, we discuss studies of the Drosophila trachea and of the vertebrate vasculature, which have revealed how new branches are formed and connect (anastomose), leading to the establishment of complex tubular networks. We briefly describe the cell behaviour underlying tracheal and vascular branching. Although similar at many levels, the branching and anastomosis processes characterized thus far show a number of differences in cell behaviour, resulting in somewhat different tube architectures in these two organs. We describe the similarities and the differences and discuss them in the context of their possible developmental significance. We finish by highlighting some old and new data, which suggest that live imaging of the development of capillary beds in adult animals might reveal yet unexplored endothelial behaviour of endothelial cells.\n\nO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=161 SRC=\"FIGDIR/small/458281_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (25K):\norg.highwire.dtl.DTLVardef@1f61f04org.highwire.dtl.DTLVardef@16e76fborg.highwire.dtl.DTLVardef@1458fd7org.highwire.dtl.DTLVardef@1e67d51_HPS_FORMAT_FIGEXP M_FIG C_FIG

developmental biology↗

bioRxiv2017

Junction-based lamellipodia drive endothelial cell rearrangements in vivo via a VE-cadherin/F-actin based oscillatory ratchet mechanism

Angiogenesis and vascular remodeling are driven by a wide range of endothelial cell behaviors, such as cell divisions, cell movements, cell shape and polarity changes. To decipher the cellular and molecular mechanism of cell movements, we have analyzed the dynamics of different junctional components during blood vessel anastomosis in vivo. We show that endothelial cell movements are associated with oscillating lamellipodia-like structures, which are orientated in the direction of these movements. These structures emerge from endothelial cell junctions and we thus call them junction-based lamellipodia (JBL). High-resolution time-lapse imaging shows that JBL are formed by F-actin based protrusions at the front end of moving cells. These protrusions also contain diffusely distributed VE-cadherin, whereas the junctional protein ZO-1 (Zona occludens 1) remains at the junction. Subsequently, a new junction is formed at the front of the JBL and the proximal junction is pulled towards the newly established distal junction. JBL function is highly dependent on F-actin dynamics. Inhibition of F-actin polymerization prevents JBL formation, whereas Rac-1 inhibition interferes with JBL oscillations. Both interventions disrupt endothelial junction formation and cell elongation. To examine the role of VE-cadherin (encoded by cdh5 gene) in this process, we generated a targeted mutation in VE-cadherin gene (cdh5ubs25), which prevents VE-cad/F-actin interaction. Although homozygous ve-cadherin mutants form JBL, these JBL are less dynamic and do not promote endothelial cell elongation. Taken together, our observations suggest a novel oscillating ratchet-like mechanism, which is used by endothelial cells to move along or over each other and thus provides the physical means for cell rearrangements.

developmental biology↗