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          Institute: MPI für Herz- und Lungenforschung (W. G. Kerckhoff Institut)     Collection: Yearbook_2014     Display Documents

ID: 682183.0, MPI für Herz- und Lungenforschung (W. G. Kerckhoff Institut) / Yearbook_2014
The polarity protein Scrib is essential for directed endothelial cell migration
Authors:Michaelis, U. R.; Chavakis, E.; Kruse, C.; Jungblut, B.; Kaluza, D.; Wandzioch, K.; Manavski, Y.; Heide, H.; Santoni, M. J.; Potente, M.; Eble, J. A.; Borg, J. P.; Brandes, R. P.
Date of Publication (YYYY-MM-DD):2013-03-15
Journal Abbrev.:Circ Res
Issue / Number:6
Start Page:924
End Page:934
Audience:Not Specified
Abstract / Description:RATIONALE: Polarity proteins are involved in the apico-basal orientation of epithelial cells, but relatively little is known regarding their function in mesenchymal cells. OBJECTIVE: We hypothesized that polarity proteins also contribute to endothelial processes like angiogenesis. METHODS AND RESULTS: Screening of endothelial cells revealed high expression of the polarity protein Scribble (Scrib). On fibronectin-coated carriers Scrib siRNA (siScrib) blocked directed but not random migration of human umbilical vein endothelial cells and led to an increased number and disturbed orientation of cellular lamellipodia. Coimmunoprecipitation/mass spectrometry and glutathione S-transferase (GST) pulldown assays identified integrin alpha5 as a novel Scrib interacting protein. By total internal reflection fluorescence (TIRF) microscopy, Scrib and integrin alpha5 colocalize at the basal plasma membrane of endothelial cells. Western blot and fluorescence activated cell sorting (FACS) analysis revealed that silencing of Scrib reduced the protein amount and surface expression of integrin alpha5 whereas surface expression of integrin alphaV was unaffected. Moreover, in contrast to fibronectin, the ligand of integrin alpha5, directional migration on collagen mediated by collagen-binding integrins was unaffected by siScrib. Mechanistically, Scrib supported integrin alpha5 recycling and protein stability by blocking its interaction with Rab7a, its translocation into lysosomes, and its subsequent degradation by pepstatin-sensitive proteases. In siScrib-treated cells, reinduction of the wild-type protein but not of PSD95, Dlg, ZO-1 (PDZ), or leucine rich repeat domain mutants restored integrin alpha5 abundance and directional cell migration. The downregulation of Scrib function in Tg(kdrl:EGFP)(s843) transgenic zebrafish embryos delayed the angiogenesis of intersegmental vessels. Conclusions: Scrib is a novel regulator of integrin alpha5 turnover and sorting, which is required for oriented cell migration and sprouting angiogenesis.
Free Keywords:Animals; Cell Migration Assays; Cell Movement/drug effects/*physiology; Cell Polarity/*physiology; Endothelial Cells/physiology; Human Umbilical Vein Endothelial Cells/*physiology; Humans; Integrin alpha5/*metabolism; Integrin alphaV/metabolism; Membrane Proteins/antagonists & inhibitors/*physiology; Mice; Neovascularization, Physiologic/*physiology; RNA, Small Interfering/pharmacology; Tumor Suppressor Proteins/antagonists & inhibitors/*physiology
External Publication Status:published
Document Type:Article
Communicated by:Svea Hümmer
Affiliations:MPI für physiologische und klinische Forschung
External Affiliations:%W NLM %G eng
Identifiers:ISSN:0009-7330 %R 10.1161/circresaha.112.300592
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