title: RhoJ/TCL regulates endothelial motility and tube formation and modulates actomyosin contractility and focal adhesion numbers.
abstract: OBJECTIVE
  RhoJ/TCL was identified by our group as an endothelial-expressed Rho GTPase. The aim of this study was to determine its tissue distribution, subcellular localization, and function in endothelial migration and tube formation.
  
  
  RESULTS
  Using in situ hybridization, RhoJ was localized to endothelial cells in a set of normal and cancerous tissues and in the vasculature of mouse embryos; endogenous RhoJ was localized to focal adhesions by immunofluorescence. The proangiogenic factor vascular endothelial growth factor activated RhoJ in endothelial cells. Using either small interfering (si)RNA-mediated knockdown of RhoJ expression or overexpression of constitutively active RhoJ (daRhoJ), RhoJ was found to positively regulate endothelial motility and tubule formation. Downregulating RhoJ expression increased focal adhesions and stress fibers in migrating cells, whereas daRhoJ overexpression resulted in the converse. RhoJ downregulation resulted in increased contraction of a collagen gel and increased phospho-myosin light chain, indicative of increased actomyosin contractility. Pharmacological inhibition of Rho-kinase (which phosphorylates myosin light chain) or nonmuscle myosin II reversed the defective tube formation and migration of RhoJ knockdown cells.
  
  
  CONCLUSIONS
  RhoJ is endothelial-expressed in vivo, activated by vascular endothelial growth factor, localizes to focal adhesions, regulates endothelial cell migration and tube formation, and modulates actomyosin contractility and focal adhesion numbers.
authors.affiliation: CRUK Molecular Angiogenesis Laboratory, Institute for Biomedical Research, The Medical School, University of Birmingham, Birmingham, United Kingdom.
authors.forename: Sukhbir
authors.initials: S
authors.lastname: Kaur
authors.forename: Katarzyna
authors.initials: K
authors.lastname: Leszczynska
authors.forename: Sabu
authors.initials: S
authors.lastname: Abraham
authors.forename: Margherita
authors.initials: M
authors.lastname: Scarcia
authors.forename: Sabina
authors.initials: S
authors.lastname: Hiltbrunner
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Marshall
authors.forename: Georgia
authors.initials: G
authors.lastname: Mavria
authors.forename: Roy
authors.initials: R
authors.lastname: Bicknell
authors.forename: Victoria L
authors.initials: VL
authors.lastname: Heath
mesh_terms: D000205:Actomyosin / Q000378:metabolism*; D000818:Animals; D002465:Cell Movement / Q000187:drug effects; D048430:Cell Shape; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000187:drug effects / Q000201:enzymology*; D005455:Fluorescent Antibody Technique; D022001:Focal Adhesions / Q000378:metabolism*; D020558:GTP Phosphohydrolases / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D017403:In Situ Hybridization; D051379:Mice; D018994:Myosin Light Chains / Q000378:metabolism; D018919:Neovascularization, Physiologic / Q000187:drug effects; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D034622:RNA Interference; D022502:Stress Fibers / Q000378:metabolism; D014162:Transfection; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism; D020741:rho GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: D018994:Myosin Light Chains; D047428:Protein Kinase Inhibitors; C485944:Rhoj protein, mouse; D042461:Vascular Endothelial Growth Factor A; D000205:Actomyosin; D054460:rho-Associated Kinases; D020558:GTP Phosphohydrolases; C417188:RHOJ protein, human; D020741:rho GTP-Binding Proteins