title: Phosphorylation of DEP-1/PTPRJ on threonine 1318 regulates Src activation and endothelial cell permeability induced by vascular endothelial growth factor.
abstract: The protein tyrosine phosphatase DEP-1/PTPRJ positively regulates Src family kinases and critical biological functions in endothelial and hematopoietic cells. Phosphorylation of DEP-1 on Y1311/Y1320 mediates the association and activation of Src, and promotes Src-dependent angiogenic responses including endothelial cell permeability. We have identified T1318 as a phosphorylated residue proximal to Y1320. The aim of this study was to determine if T1318 phosphorylation exerts a regulatory role over the function of DEP-1. We show that phosphorylation of DEP-1 on Y1320 was reduced when T1318 was mutated. This led to the decreased association of DEP-1 T1318A with Src, and defective Src activation in both HEK 293T and VEGF-stimulated endothelial cells. Consistent with these findings, VEGF-induced tyrosine phosphorylation of VE-cadherin, its association to β-arrestin1/2, and cell permeability were impaired in cells expressing DEP-1 T1318A. Conversely, expression of the phosphomimetic mutant DEP-1 T1318E constitutively enhanced the phosphorylation of Y1320 and VE-cadherin over that induced by WT DEP-1, and resulted in increased VEGF-dependent permeability. DEP-1 T1318 is part of a CK2 consensus phosphorylation site and was identified as a CK2 substrate. Modulation of CK2 expression or activity in endothelial cells regulated T1318 phosphorylation, and correlated with the status of Y1320 phosphorylation, Src activation, and cell permeability. CK2-dependent phosphorylation of DEP-1 T1318 promotes Y1320 phosphorylation and Src activation upon VEGF stimulation. Phosphorylation of T1318 is thus part of a regulatory mechanism that channels the activity of DEP-1 towards Src to allow its optimal activation and the promotion of endothelial cell permeability.
authors.affiliation: CRCHUM - Centre de recherche du Centre Hospitalier de l'Université de Montréal and Institut du Cancer de Montréal, Montréal, Québec, Canada.
authors.forename: Kathleen
authors.initials: K
authors.lastname: Spring
authors.affiliation: CRCHUM - Centre de recherche du Centre Hospitalier de l'Université de Montréal and Institut du Cancer de Montréal, Montréal, Québec, Canada.
authors.forename: Line
authors.initials: L
authors.lastname: Lapointe
authors.affiliation: CRCHUM - Centre de recherche du Centre Hospitalier de l'Université de Montréal and Institut du Cancer de Montréal, Montréal, Québec, Canada.
authors.forename: Christine
authors.initials: C
authors.lastname: Caron
authors.affiliation: CRCHUM - Centre de recherche du Centre Hospitalier de l'Université de Montréal and Institut du Cancer de Montréal, Montréal, Québec, Canada.
authors.forename: Simon
authors.initials: S
authors.lastname: Langlois
authors.affiliation: CRCHUM - Centre de recherche du Centre Hospitalier de l'Université de Montréal and Institut du Cancer de Montréal, Montréal, Québec, Canada; Department of Medicine, Université de Montréal, Montréal, Québec, Canada. Electronic address: isabelle.royal@umontreal.ca.
authors.forename: Isabelle
authors.initials: I
authors.lastname: Royal
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D047390:Casein Kinase II / Q000378:metabolism; D002417:Cattle; D002463:Cell Membrane Permeability; D004789:Enzyme Activation; D057809:HEK293 Cells; D061307:Human Umbilical Vein Endothelial Cells / Q000378:metabolism*; D006801:Humans; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D054631:Receptor-Like Protein Tyrosine Phosphatases, Class 3 / Q000378:metabolism; D013912:Threonine / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A / Q000502:physiology*; D019061:src-Family Kinases / Q000378:metabolism
chemical_list: C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D013912:Threonine; D019061:src-Family Kinases; D047390:Casein Kinase II; C486288:PTPRJ protein, human; D054631:Receptor-Like Protein Tyrosine Phosphatases, Class 3
keywords: Protein tyrosine phosphatase; Src; VE-cadherin; Vascular endothelial growth factor; Vascular permeability