Genetic Analysis of Blood Vessel Morphogenesis: the role of Rasip1 in the regulation of junctional dynamics during angiogenic sprouting and lumen formation

Lee, Minkyoung. Genetic Analysis of Blood Vessel Morphogenesis: the role of Rasip1 in the regulation of junctional dynamics during angiogenic sprouting and lumen formation. 2021, Doctoral Thesis, University of Basel, Faculty of Science.


Official URL: https://edoc.unibas.ch/85048/

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The formation of new blood vessels from pre-existing ones, or angiogenesis, is a critical morphogenetic process during embryo development. Angiogenesis also happens in the adult body during wound healing and in pathophysiological conditions such as in cancer growth. Blood vessel morphogenesis heavily relies on the cell-cell interactions, which enable endothelial cells (ECs) to rearrange and change their shape in order to extend the nascent sprout and to form a multi- cellular tube. At the cellular level, the formation of a vascular tube is driven by a collection of dynamic EC behaviors including cell migration, cell shape changes and cell rearrangements. Here, I show that such cell interactions require dynamic VE (vascular endothelial)-cadherin (Cdh5) and an F-actin oscillatory machinery. Specifically, I show that in zebrafish mutant embryos which lack the VE-cadherin actin-binding site, elongation of cell-cell junctions and cellular rearrangements during anastomosis are perturbed. Therefore, this mutant analysis highlights the collaborative interaction between F-actin and VE-cadherin to generate mechanical forces between two cells, an important aspect for cell shape changes that ultimately leads to multicellular vessel formation.
To gain more insight into the in vivo regulation of morphogenetic junctional dynamics, I analyzed mutants for the zebrafish ras-interacting protein 1 (rasip1), a putative regulator of EC junctional dynamics via VE-cadherin interactions and ras-associating and dilute domain-containing protein (radil), a protein closely related to Rasip1 involving RhoA/ROCK signaling. The analysis of rasip1/ radilb single and double mutants reveals redundant and distinct functions of these genes during vascular morphogenesis. Whilst both proteins are required for cell shape regulation and luminal stability, Rasip1 is unique in its role for the organization of EC junctions at the cell-cell interfaces during angiogenic sprouting and anastomosis. The aberrant localization of junctional molecules (such as VE-cadherin, ZO1, PECAM1 and ESAMa) in rasip1 mutants induces remodeling defects. In particular, this mis-localization leads to loss of junctional clearance, reduction of junctional elongation and perturbation of actin localization at the junctions, thereby preventing lumen formation during development of multicellular vessels. I have also employed loss-of-function analysis to pinpoint the molecular pathways, which underlie the respectivefunctions of Rasip1 and Radil. Specifically, Ccm1 and Heg1 are known to localize the Rasip1 protein at apical sites/membranes as well as to facilitate interaction between Rasip1 and VE-cadherin. Knockdown of either of them shows similar mis-localization patterning and destabilization of junctions as the rasip1 mutant.
Taken together, my observations suggest a mechanism in which Rasip1/Radil act at the junctional/cortical F-actin interface thereby coupling junctional remodeling with dynamic cell shape changes. This study demonstrates that the molecular pathway of the Rasip1-Ccm1-Heg1 regulates the in vivo clearance of adherens junction proteins, thereby resulting in the formation of the ring-shaped junction formation and mediating cell elongation and lumen tubulogenesis during vascular development.
Advisors:Affolter, Markus and Vermot, Julien
Faculties and Departments:05 Faculty of Science > Departement Biozentrum > Growth & Development > Cell Biology (Affolter)
UniBasel Contributors:Affolter, Markus
Item Type:Thesis
Thesis Subtype:Doctoral Thesis
Thesis no:14521
Thesis status:Complete
Number of Pages:141
Identification Number:
  • urn: urn:nbn:ch:bel-bau-diss145216
edoc DOI:
Last Modified:10 Dec 2021 08:44
Deposited On:10 Dec 2021 08:44

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